Assay for vapor condensates

ABSTRACT

The present invention relates to provide, among other things, the methods, devices, and systems that can simply and quickly collecting and analyzing a tiny amount of vapor condensates (e.g. exhaled breath condensate (EBC)).

CROSS-REFERENCE TO RELATED APPLICATIONS

This application is continuation application of U.S. Non-provisionalapplication Ser. No. 16/485,126, filed on Aug. 9, 2019, which is a § 371national stage application of International ApplicationPCT/US2018/018520 filed on Feb. 16, 2018, which claims the benefit ofpriority to U.S. Provisional Application (“U.S.PA” hereinafter) No.62/460,088, filed on Feb. 16, 2017, U.S.PA No. 62/460,091, filed on Feb.16, 2017, U.S.PA No. 62/460,083, filed on Feb. 16, 2017, U.S.PA No.62/460,076, filed on Feb. 16, 2017, U.S.PA No. 62/460,075, filed on Feb.16, 2017, U.S.PA No. 62/460,069, filed on Feb. 16, 2017, U.S.PA No.62/460,062, filed on Feb. 16, 2017, U.S.PA No. 62/460,047, filed on Feb.16, 2017, U.S.PA No. 62/459,972, filed on Feb. 16, 2017, U.S.PA No.62/459,920, filed on Feb. 16, 2017, PCT Application No. PCT/US18/18405,filed on Feb. 15, 2018, PCT Application No. PCT/US18/18108, filed onFeb. 14, 2018, PCT Application No. PCT/US18/18007, filed on Feb. 13,2018, PCT Application No. PCT/US18/17716, filed on Feb. 9, 2018, PCTApplication No. PCT/US18/17713, filed on Feb. 9, 2018, PCT ApplicationNo. PCT/US18/17712, filed on Feb. 9, 2018, PCT Application No.PCT/US18/17504, filed on Feb. 8, 2018, PCT Application No.PCT/US18/17501, filed on Feb. 8, 2018, PCT Application No.PCT/US18/17499, filed on Feb. 8, 2018, PCT Application No.PCT/US18/17489, filed on Feb. 8, 2018, PCT Application No.PCT/US18/17492, filed on Feb. 8, 2018, PCT Application No.PCT/US18/17494, filed on Feb. 8, 2018, PCT Application No.PCT/US18/17502, filed on Feb. 8, 2018, and PCT Application No.PCT/US18/17307, filed on Feb. 7, 2018, the contents of which are reliedupon and incorporated herein by reference in their entirety. The entiredisclosure of any publication or patent document mentioned herein isentirely incorporated by reference.

FIELD

The present invention is related to the field of bio/chemical sampling,sensing, assays and applications.

BACKGROUND

In bio/chemical vapor condensate sample analysis, particularly exhaledbreath condensate (EBC), there is a need for methods and devices thatcan simplify the sample collection and measurement processes, that canaccelerate the process (e.g. binding, mixing reagents, etc.) andquantify the parameters (e.g. analyte concentration, the sample volume,etc.), that can handle samples with small volume, that allow an entireassay performed in less than a minute, that allow an assay performed bya smartphone (e.g. mobile phone), that allow non-professional to performan assay her/himself, and that allow a test result to be communicatedlocally, remotely, or wirelessly to different relevant parties.

The present invention relates to provide, among other things, themethods, devices, and systems that can address these needs.

SUMMARY OF INVENTION

The following brief summary is not intended to include all features andaspects of the present invention.

The present invention is related to provide, among other things, thedevice, apparatus, systems, and methods for: (i) rapidly and simplycollecting a tiny amount (a volume as small as 10 fL (femto-Liter) in asingle drop) of a vapor condensate sample (e.g. the exhaled breathcondensate (EBC) from a subject), (ii) preventing or significantlyreducing an evaporation of the collected vapor condensate sample, (iii)analyzing the sample, analyzing the sample by mobile-phone, (iv) andperforming such collection and analysis by a person without anyprofessionals; (v) the devices can get the sample thickness informationeither using spacers or without a spacers.

Since the exhaled breath condensate (EBC) and other vapor condensateshare many common properties, the disclosure uses EBC as arepresentative to illustrate certain embodiments of the presentinvention, but such presentation should not be construed as anylimitations of the present invention.

BRIEF DESCRIPTION OF THE DRAWINGS

The skilled artisan will understand that the drawings, described below,are for illustration purposes only. The drawings are not intended tolimit the scope of the present teachings in any way. The drawings maynot be in scale. In the figures that present experimental data points,the lines that connect the data points are for guiding a viewing of thedata only and have no other means.

FIG. 1 . An illustration of certain aspects of an exemplary device andmethods of collecting exhaled breath condensate (EBC) using a SiEBCA(Single-drop EBC Collector/Analyzer), that comprises a pair of movableplates without using spacers.

FIG. 2 . An illustration of certain aspects of an exemplary device andmethods of collecting exhaled breath condensate (EBC) using a SiEBCA(Single-drop EBC Collector/Analyzer).

FIG. 3 . An illustration of different formations of EBC at closedconfiguration of SiEBCA depends on spacer height. In closedconfiguration-1: If spacer height is smaller than average height of EBCat open configuration; at closed configuration, EBC become a continuousthin film contacting both collection and cover plates and may have airisolated pockets. In the closed configuration-2: If spacer height islarger than average height of EBC at open configuration; at closedconfiguration EBC become isolated puddle(s) that contact both collectionand cover plates, and that are larger but fewer than that at the openconfiguration.

FIG. 4 . An illustration of an embodiment of the devices and the methodsof a SiEBCA (Single-drop EBC Collector/Analyzer).

FIG. 5 a An illustration of a SiEBCA with both “open spacer” and“enclosed spacer”, where the open spacer is a post (pillar) while theenclosed spacer is a ring spacer (4) and a four-chamber grid spacer (5).FIG. 5 b shows reducing binding or mixing time by reducing the samplethickness using two pates, spacers, and compression (shown incross-section). Panel (1) illustrates reducing the time for bindingentities in a sample to a binding site on a solid surface (X-(Volume toSurface)). Panel (2) illustrates reducing the time for binding entities(e.g. reagent) stored on a surface of plate to a binding site on asurface of another surface (X-(Surface to Surface)). Panel (3)illustrates reducing the time for adding reagents stored on a surface ofa plate into a sample that is sandwiched between the plate and otherplate (X-(Surface to Volume)).

FIG. 6 schematically illustrates the optical measurement and imagingtaken for the measurement of the EBC sample thickness and lateral area,respectively.

FIG. 7 demonstrates the principle of plate spacing measurement based onF-P cavity resonance.

FIG. 8 shows microscopic images of the EBC sample collected using theexemplary SiEBCA device without spacers.

DETAILED DESCRIPTION OF EXEMPLARY EMBODIMENTS

The following detailed description illustrates some embodiments of theinvention by way of example and not by way of limitation. If any, thesection headings and any subtitles used herein are for organizationalpurposes only and are not to be construed as limiting the subject matterdescribed in any way. The contents under a section heading and/orsubtitle are not limited to the section heading and/or subtitle, butapply to the entire description of the present invention.

The citation of any publication is for its disclosure prior to thefiling date and should not be construed as an admission that the presentclaims are not entitled to antedate such publication by virtue of priorinvention. Further, the dates of publication provided can be differentfrom the actual publication dates which can need to be independentlyconfirmed.

Definitions

Unless defined otherwise, all technical and scientific terms used hereinhave the same meaning as commonly understood by one of ordinary skill inthe art to which this disclosure belongs. Although any methods andmaterials similar or equivalent to those described herein can also beused in the practice or testing of the present teachings, some exemplarymethods and materials are now described.

The terms “exhaled breath condensate (EBC)” and “vapor condensate (VC)”are, unless specifically stated, interchangeable.

The term “SiEBCA” and “SiEBC” are interchangeable.

The terms “polynucleotide”, “nucleotide”, “nucleotide sequence”,“nucleic acid”, “nucleic acid molecule”, “nucleic acid sequence” and“oligonucleotide” are used interchangeably, and can also include pluralsof each respectively depending on the context in which the terms areutilized. The term “capture agent” as used herein, refers to a bindingmember, e.g. nucleic acid molecule, polypeptide molecule, or any othermolecule or compound, that can specifically bind to its binding partner,e.g., a second nucleic acid molecule containing nucleotide sequencescomplementary to a first nucleic acid molecule, an antibody thatspecifically recognizes an antigen, an antigen specifically recognizedby an antibody, a nucleic acid aptamer that can specifically bind to atarget molecule, etc.

The term “a secondary capture agent” which can also be referred to as a“detection agent” refers a group of biomolecules or chemical compoundsthat have highly specific affinity to the antigen. The secondary captureagent can be strongly linked to an optical detectable label, e.g.,enzyme, fluorescence label, or can itself be detected by anotherdetection agent that is linked to an optical detectable label throughbioconjugation (Hermanson, “Bioconjugate Techniques” Academic Press, 2ndEd., 2008).

The term “capture agent-reactive group” refers to a moiety of chemicalfunction in a molecule that is reactive with capture agents, i.e., canreact with a moiety (e.g., a hydroxyl, sulfhydryl, carboxyl or aminegroup) in a capture agent to produce a stable strong, e.g., covalentbond.

The terms “specific binding” and “selective binding” refer to theability of a capture agent to preferentially bind to a particular targetanalyte that is present in a heterogeneous mixture of different targetanalytes. A specific or selective binding interaction will discriminatebetween desirable (e.g., active) and undesirable (e.g., inactive) targetanalytes in a sample, typically more than about 10 to 100-fold or more(e.g., more than about 1000- or 10,000-fold).

The term “analyte” refers to a molecule (e.g., a protein, peptides, DNA,RNA, nucleic acid, or other molecule), cells, tissues, viruses, andnanoparticles with different shapes.

The term “assaying” refers to testing a sample to detect the presenceand/or abundance of an analyte.

As used herein, the terms “determining,” “measuring,” and “assessing,”and “assaying” are used interchangeably and include both quantitativeand qualitative determinations.

As used herein, the term “light-emitting label” refers to a label thatcan emit light when under an external excitation. This can beluminescence. Fluorescent labels (which include dye molecules or quantumdots), and luminescent labels (e.g., electro- or chemi-luminescentlabels) are types of light-emitting label. The external excitation islight (photons) for fluorescence, electrical current forelectroluminescence and chemical reaction for chemi-luminescence. Anexternal excitation can be a combination of the above.

The phrase “labeled analyte” refers to an analyte that is detectablylabeled with a light emitting label such that the analyte can bedetected by assessing the presence of the label. A labeled analyte maybe labeled directly (i.e., the analyte itself may be directly conjugatedto a label, e.g., via a strong bond, e.g., a covalent or non-covalentbond), or a labeled analyte may be labeled indirectly (i.e., the analyteis bound by a secondary capture agent that is directly labeled).

The terms “hybridizing” and “binding”, with respect to nucleic acids,are used interchangeably.

The term “capture agent/analyte complex” is a complex that results fromthe specific binding of a capture agent with an analyte. A capture agentand an analyte for the capture agent will usually specifically bind toeach other under “specific binding conditions” or “conditions suitablefor specific binding”, where such conditions are those conditions (interms of salt concentration, pH, detergent, protein concentration,temperature, etc.) which allow for binding to occur between captureagents and analytes to bind in solution. Such conditions, particularlywith respect to antibodies and their antigens and nucleic acidhybridization are well known in the art (see, e.g., Harlow and Lane(Antibodies: A Laboratory Manual Cold Spring Harbor Laboratory, ColdSpring Harbor, N.Y. (1989) and Ausubel, et al, Short Protocols inMolecular Biology, 5th ed., Wiley & Sons, 2002).

A subject may be any human or non-human animal. A subject may be aperson performing the instant method, a patient, a customer in a testingcenter, etc.

As used herein, a “diagnostic sample” refers to any biological samplethat is a bodily byproduct, such as bodily fluids, that has been derivedfrom a subject. The diagnostic sample may be obtained directly from thesubject in the form of liquid, or may be derived from the subject byfirst placing the bodily byproduct in a solution, such as a buffer.Exemplary diagnostic samples include, but are not limited to, saliva,serum, blood, sputum, urine, sweat, lacrima, semen, feces, breath,biopsies, mucus, etc.

As used herein, an “environmental sample” refers to any sample that isobtained from the environment. An environmental sample may includeliquid samples from a river, lake, pond, ocean, glaciers, icebergs,rain, snow, sewage, reservoirs, tap water, drinking water, etc.; solidsamples from soil, compost, sand, rocks, concrete, wood, brick, sewage,etc.; and gaseous samples from the air, underwater heat vents,industrial exhaust, vehicular exhaust, etc. Typically, samples that arenot in liquid form are converted to liquid form before analyzing thesample with the present method.

As used herein, a “foodstuff sample” refers to any sample that issuitable for animal consumption, e.g., human consumption. A foodstuffsample may include raw ingredients, cooked food, plant and animalsources of food, preprocessed food as well as partially or fullyprocessed food, etc. Typically, samples that are not in liquid form areconverted to liquid form before analyzing the sample with the presentmethod.

The term “diagnostic,” as used herein, refers to the use of a method oran analyte for identifying, predicting the outcome of and/or predictingtreatment response of a disease or condition of interest. A diagnosismay include predicting the likelihood of or a predisposition to having adisease or condition, estimating the severity of a disease or condition,determining the risk of progression in a disease or condition, assessingthe clinical response to a treatment, and/or predicting the response totreatment.

A “biomarker,” as used herein, is any molecule or compound that is foundin a sample of interest and that is known to be diagnostic of orassociated with the presence of or a predisposition to a disease orcondition of interest in the subject from which the sample is derived.Biomarkers include, but are not limited to, polypeptides or a complexthereof (e.g., antigen, antibody), nucleic acids (e.g., DNA, miRNA,mRNA), drug metabolites, lipids, carbohydrates, hormones, vitamins,etc., that are known to be associated with a disease or condition ofinterest.

A “condition” as used herein with respect to diagnosing a healthcondition, refers to a physiological state of mind or body that isdistinguishable from other physiological states. A health condition maynot be diagnosed as a disease in some cases. Exemplary health conditionsof interest include, but are not limited to, nutritional health; aging;exposure to environmental toxins, pesticides, herbicides, synthetichormone analogs; pregnancy; menopause; andropause; sleep; stress;prediabetes; exercise; fatigue; chemical balance; etc.

The term “entity” refers to, but not limited to proteins, peptides, DNA,RNA, nucleic acid, molecules (small or large), cells, tissues, viruses,nanoparticles with different shapes, that would bind to a “bindingsite”. The entity includes the capture agent, detection agent, andblocking agent.

The “entity” includes the “analyte”, and the two terms are usedinterchangeably.

The term “binding site” refers to a location on a solid surface that canimmobilize an entity in a sample.

The term “entity partners” refers to, but not limited to proteins,peptides, DNA, RNA, nucleic acid, molecules (small or large), cells,tissues, viruses, nanoparticles with different shapes, that are on a“binding site” and would bind to the entity. The entity, include, butnot limited to, capture agents, detection agents, secondary detectionagents, or “capture agent/analyte complex”.

The term “smart phone” or “mobile phone”, which are usedinterchangeably, refers to the type of phones that has a camera andcommunication hardware and software that can take an image using thecamera, manipulate the image taken by the camera, and communicate datato a remote place. In some embodiments, the Smart Phone has a flashlight.

The term “average linear dimension” of an area is defined as a lengththat equals to the area times 4 then divided by the perimeter of thearea. For example, the area is a rectangle, that has width w, and lengthL, then the average of the linear dimension of the rectangle is4*W*L/(2*(L+W)) (where “*” means multiply and “/” means divide). By thisdefinition, the average line dimension is, respectively, W for a squareof a width W, and d for a circle with a diameter d. The area includes,but not limited to, the area of a binding site or a storage site.

The term “period” of periodic structure array refers to the distancefrom the center of a structure to the center of the nearest neighboringidentical structure.

The term “storage site” refers to a site of an area on a plate, whereinthe site contains reagents to be added into a sample, and the reagentsare capable of being dissolving into the sample that is in contract withthe reagents and diffusing in the sample.

The term “relevant” means that it is relevant to detection of analytes,quantification and/or control of analyte or entity in a sample or on aplate, or quantification or control of reagent to be added to a sampleor a plate.

The term “hydrophilic”, “wetting”, or “wet” of a surface means that thecontact angle of a sample on the surface is less than 90 degree.

The term “hydrophobic”, “non-wetting”, or “does not wet” of a surfacemeans that the contact angle of a sample on the surface is equal to orlarger than 90 degree.

The term “variation” of a quantity refers to the difference between theactual value and the desired value or the average of the quantity. Andthe term “relative variation” of a quantity refers to the ratio of thevariation to the desired value or the average of the quantity. Forexample, if the desired value of a quantity is Q and the actual value is(Q+ □), then the □/(Q+□) is the relative variation. The term “relativesample thickness variation” refers to the ratio of the sample thicknessvariation to the average sample thickness.

The term “optical transparent” refers to a material that allows atransmission of an optical signal, wherein the term “optical signal”refers to, unless specified otherwise, the optical signal that is usedto probe a property of the sample, the plate, the spacers, thescale-marks, any structures used, or any combinations of thereof.

The term “none-sample-volume” refers to, at a closed configuration of aCROF process, the volume between the plates that is occupied not by thesample but by other objects that are not the sample. The objectsinclude, but not limited to, spacers, air bubbles, dusts, or anycombinations of thereof. Often none-sample-volume(s) is mixed inside thesample.

The term “saturation incubation time” refers to the time needed for thebinding between two types of molecules (e.g. capture agents andanalytes) to reach an equilibrium. For a surface immobilization assay,the “saturation incubation time” refers the time needed for the bindingbetween the target analyte (entity) in the sample and the binding siteon plate surface reaches an equilibrium, namely, the time after whichthe average number of the target molecules (the entity) captured andimmobilized by the binding site is statistically nearly constant.

In some cases, the “analyte” and “binding entity” and “entity” areinterchangeable.

The term “first plate” and “collection plate are interchangeable. Theterm “second plate” and “cover plate” are interchangeable.

A “processor,” “communication device,” “mobile device,” refer tocomputer systems that contain basic electronic elements (including oneor more of a memory, input-output interface, central processing unit,instructions, network interface, power source, etc.) to performcomputational tasks. The computer system may be a general purposecomputer that contains instructions to perform a specific task, or maybe a special-purpose computer.

A “site” or “location” as used in describing signal or datacommunication refers to the local area in which a device or subjectresides. A site may refer to a room within a building structure, such asa hospital, or a smaller geographically defined area within a largergeographically defined area. A remote site or remote location, withreference to a first site that is remote from a second site, is a firstsite that is physically separated from the second site by distanceand/or by physical obstruction. The remote site may be a first site thatis in a separate room from the second site in a building structure, afirst site that is in a different building structure from the secondsite, a first site that is in a different city from the second site,etc.

As used herein, the term “sample collection site” refers to a locationat which a sample may be obtained from a subject. A sample collectionsite may be, for example, a retailer location (e.g., a chain store,pharmacy, supermarket, or department store), a provider office, aphysician's office, a hospital, the subject's home, a military site, anemployer site, or other site or combination of sites. As used herein,the term “sample collection site” may also refer to a proprietor orrepresentative of a business, service, or institution located at, oraffiliated with, the site.

As used herein, “raw data” includes signals and direct read-outs fromsensors, cameras, and other components and instruments which detect ormeasure properties or characteristics of a sample.

“Process management,” as used herein, refers to any number of methodsand systems for planning and/or monitoring the performance of a process,such as a sample analysis process.

One with skill in the art will appreciate that the present invention isnot limited in its application to the details of construction, thearrangements of components, category selections, weightings,pre-determined signal limits, or the steps set forth in the descriptionor drawings herein. The invention is capable of other embodiments and ofbeing practiced or being carried out in many different ways.

1. Vapor Condensate Assay by Two Movable Plates without Using SpacersOne aspect of the present invention is that we developed andexperimentally demonstrated of collecting and assaying vapor condensate(VC) (e.g. EBC samples) using a pair of movable plates without usingspacers.

Exhaled breath condensate (EBC) analysis is a noninvasive method ofdetecting biomarkers, mainly coming from the lower respiratory tract.EBC is collected during quiet breathing, as a product of cooling andcondensation of the exhaled aerosol.

An exemplary device and method of collecting exhaled breath condensate(EBC) using a SiEBC (single-drop exhaled breath condensatecollector/analyzer) or SiVC (single-drop vapor condensatecollector/analyzer) that does not have spacers, as illustrated in FIG. 1, comprises the basic steps:

(1) exhaling breath onto the collection plate (FIG. 1-1 ). A subject(e.g. human) breathe onto a plate, termed “collection plate”, and thebreath condenses into EBC, which are in droplets with different sizes,depending on the breathing time. For a short breathing time mostdroplets are separated from each other. The surface of the collectionplate that collects the EBC is termed the sample surface;

(2) placing a cover plate over the collection plate and pressing themtogether (FIG. 1-2 ). A cover plate with spacers (which are used forregulating the spacing between the cover plate and the substrate plate)is placed on top of the sample surface; and

(3) pressing plates into a “Closed-Configuration (FIG. 1-3 ). The coverplate and the substrate are compressed together with at least a part ofthe EBC between the plates.

In the method of FIG. 1 , the initial droplets are pressed into a thinlayer EBC of a thickness that is regulated by the plates and spacers(not shown).

One reason for using the wording of “single drop” in SiEBCA is that inprinciple, the SiEBCA can detect and analyze a single drop of EBCdeposited on the plate.

In the description of the present invention, “the substrate plate” and“the cover plate” are respectively interchangeable with “the firstplate” and “the second plate”.

In some embodiments, the plates are cooled to reduce the evaporation ofcollected EBC.

-   AA0-1. A device for collecting and analyzing vapor condensate (VC)    sample, comprising:

a collection plate and a cover plate, wherein:

-   -   i. the plates are movable relative to each other into different        configurations;    -   ii. one or both plates are flexible; and    -   iii. each of the plates has, on its inner respective surface, a        sample contact area for contacting a vapor condensate (VC)        sample that contains an analyte;    -   wherein one of the configurations is an open configuration, in        which: the two plates are either completely or partially        separated apart, and the VC sample is deposited on one or both        of the plates; and    -   wherein another of the configurations is a closed configuration        which is configured after the VC sample deposition in the open        configuration; and in the closed configuration: at least a part        of the VC sample is between the two plates and in contact with        the two plates, and has a thickness that (a) is regulated by the        two sample contact surfaces of the plates without using spacers,        and (b) is equal to or less than 30 um with a small variation.

-   AA0-2 Another device is provided herein for collecting and analyzing    vapor condensate (VC) sample, comprising:

a collection plate and a cover plate, wherein:

-   -   i. the plates are movable relative to each other into different        configurations;    -   ii. one or both plates are flexible; and    -   iii. each of the plates has, on its respective surface, a sample        contact area for contacting a vapor condensate (VC) sample that        contains an analyte;        -   wherein one of the configurations is an open configuration,            in which: the two plates are either completely or partially            separated apart, and the VC sample is deposited on one or            both of the plates; and        -   wherein another of the configurations is a closed            configuration which is configured after the VC sample            deposition in the open configuration; and in the closed            configuration: at least a part of the VC sample is between            the two plates and in contact with the two plates, and has a            thickness that is regulated by the plate spacing without            using spacers.

In some embodiments, the thickness of the sample in a closedconfiguration of the plates is measured. The sample thickness in a plateclosed configuration include Fabry-Pérot interferometer, or differentoptical focusing methods, or others.

Experiment-1

In an experiment by the present invention, an exemplary SiEBCA devicecomprises a collection plate of 25 mm×25 mm×1 mm PMMA planar plate withuntreated surfaces, and a cover plate of 25 mm×25 mm×0.175 mm PMMAplanar plate with bare untreated surfaces. The EBC sample was collectedby having a subject breathe on a collection plate for 2 sec and a coverplate was immediately brought to cover the collection plate and pressedagainst it as described above. Later, the SiEBCA together with thesample collected therein were subject to optical measurement andmicroscopy imaging.

FIG. 6 schematically illustrates the optical measurement and imagingtaken for the measurement of the EBC sample thickness and lateral area,respectively. As shown in panel (A), Fabry-Pérot interferometer was usedto measure the F-P cavity resonance in the reflectance spectra at 25points on the 4×4 grid artificially generated in the center of theSiEBCA device, from which the plate spacing (and the sample thickness)is thus deduced. Each of the 25 measuring points is about 2 um by 2 umin area, and all 25 points cover an area of 20 mm by 20 mm. An averageplate spacing over the 25 points was taken as the estimate of the samplethickness (

). As shown in panel (B), a direct photo of the SiEBCA device was takento delineate the general contour of the EBC sample between the twoplates and measure the overall lateral area (S_(t)). Then microscopicimages were taken at each of the 25 points (each image covers an areaS_(t) of 1.6 mm×1.1 mm), and then these images were analyzed by an imageprocessing software to recognize and measure the total area of the airbubbles (Sb) in each image.

To estimate the total EBC sample lateral area, first, the percentage ofEBC liquid lateral area (a_(i)) for each measuring point is calculatedas (Si−Sb)/Si×100%; second, an average value (ã) is taken from all 25points; and finally, the total EBC sample lateral area (S_(EBC)) isestimated as S_(t)*ã.

The volume of the EBC sample (V_(EBC)) is thus determined as S_(EBC)*

.

FIG. 7 demonstrates the principle of plate spacing measurement based onF-P cavity resonance. Panel (a) shows the schematic of F-P cavity fromthe SiEBCA device; panel (b) shows the typical reflectance spectrum andresonances from the device. The plate spacing (h) at each measuringpoint is calculated as:

$h = \frac{c}{2n\Delta v}$

where h is the plate spacing, c is light speed, Δv is the period infrequency domain and n is the reflective index of the EBC liquid.

As described above, the average EBC sample thickness is equal to

$\frac{\sum_{1}^{25}h}{25}.$

EBC sample thickness uniformity is calculated as

$\sqrt{\frac{\sum_{1}^{25}\left( {h - H} \right)^{2}}{25}}.$

FIG. 8 shows microscopic images of the EBC sample collected using theexemplary SiEBCA device without spacers. Panels (a)-(b) respectivelyshow the images of the EBC samples at the closed configuration afterhand pressing the two plates with low, medium, and high pressingstrength. Low strength was less than 10 kg, high strength was higherthan 15 kg, and medium strength was in between the low and highstrength.

Under these three different conditions, the performance of the exemplarySiEBCA device without spacers was examined and summarized in Table 3,based on the measurement and calculation methods described above. Asshown in Table 3 and FIG. 8 , low strength gives thicker liquidthickness with larger bubble area, while high strength gives thinner EBCsample layer with smaller bubble area.

TABLE 3 Performance of SiEBCA without spacers Average Average EBCCollected EBC Press EBC area EBC Area S_(EBC) thickness EBC volumethickness Strength percentage ã (mm²)

 (um) V_(EBC) (uL) uniformity 1 Low 38% 240 1.45 0.35 58% 2 Medium 72%450 0.87 0.39 47% 3 High 98% 620 0.51 0.32 43%

2. Vapor Condensate Assay by Two Movable Plates with Spacers

-   AA1-1 In some embodiments, as illustrated in FIG. 1 (2), a device is    provided herein for collecting and analyzing vapor condensate (VC)    sample, comprising:

a collection plate, a cover plate, and spacers, wherein:

-   -   i. the plates are movable relative to each other into different        configurations;    -   ii. one or both plates are flexible;    -   iii. each of the plates has, on its respective inner surface, a        sample contact area for contacting a vapor condensate (VC)        sample that contains an analyte;    -   iv. the spacers are fixed to the respective inner surface of one        or both of the plates, and have a predetermined substantially        uniform height and a predetermined constant inter-spacer        distance and wherein at least one of the spacers is inside the        sample contact area;    -   wherein one of the configurations is an open configuration, in        which: the two plates are either completely or partially        separated apart, the spacing between the plates is not regulated        by the spacers, and the VC sample is deposited on one or both of        the plates; and    -   wherein another of the configurations is a closed configuration        which is configured after the VC sample deposition in the open        configuration; and in the closed configuration: at least a part        of the VC sample is between the two plates and in contact with        the two plates, and has a highly uniform thickness that is        regulated by the spacers and the two sample contact surfaces of        the plates and is equal to or less than 30 um.

3. Different Embodiments for Devices with or without Spacers

In some embodiments, the device further comprises, on one or bothplates, one or a plurality of dry binding sites and/or one or aplurality of reagent sites. In some embodiments, the sample is exhalebreath condensate.

In some embodiments, the sample is a vapor from a biological sample, anenvironmental sample, a chemical sample, or clinical sample. In someembodiments, wherein the analyte comprises a molecule (e.g., a protein,peptides, DNA, RNA, nucleic acid, or other molecules), cells, tissues,viruses, and nanoparticles with different shapes. In some embodiments,wherein the analyte comprises volatile organic compounds (VOCs). In someembodiments, wherein the analyte comprises nitrogen, oxygen, CO2, H2O,and inert gases. In some embodiments, wherein the analyte is stained.

In some embodiments, the device may comprise a dry reagent coated on oneor both of the plates. In some embodiments, the dry reagent may bind toan analyte in the blood an immobilize the analyte on a surface on one orboth of the plates. In these embodiments, the reagent may be an antibodyor other specific binding agent, for example. This dry reagent may havea pre-determined area. In other embodiments, the device may comprise areleasable dry reagent on one or more of the plates, e.g., a labeledreagent such as a cell stain or a labeled detection agent such as anantibody or the like. In some cases, there may be a release time controlmaterial on the plate that contains the releasable dry reagent, whereinthe release time control material delays the time that the releasabledry regent is released into the blood sample.

In some cases, the release time control material delays the time thatthe dry regent is released into the blood sample by at least 3 seconds,e.g., at least 5 seconds or at least 10 seconds. Some embodiments, thedrive may contain multiple dry binding sites and/or multiple reagentsites, thereby allowing multiplex assays to be performed. In some cases,the areas occupied by the drying binding sites may oppose the areasoccupied by the reagent sites when the plates are in the closedposition.

In some embodiments, the regent comprises labeling or stainingreagent(s).

In some embodiments, the spacers regulating the layer of uniformthickness (i.e., the spacers that are spacing the plates away from eachother in the layer) have a “filling factor” of at least 1%, e.g., atleast 2% or at least 5%, wherein the filling factor is the ratio of thespacer area that is in contact with the layer of uniform thickness tothe total plate area that is in contact with the layer of uniformthickness. In some embodiments, for spacers regulating the layer ofuniform thickness, the Young's modulus of the spacers times the fillingfactor of the spacers is equal or larger than 10 MPa, e.g., at least 15MPa or at least 20 MPa, where the filling factor is the ratio of thespacer area that is in contact with the layer of uniform thickness tothe total plate area that is in contact with the layer of uniformthickness. In some embodiments, the thickness of the flexible platetimes the Young's modulus of the flexible plate is in the range 60 to750 GPa-um, e.g., 100 to 300 GPa-um, 300 to 550 GPa-um, or 550 to 750GPa-um. In some embodiments, for a flexible plate, the fourth power ofthe inter-spacer-distance (ISD) divided by the thickness of the flexibleplate (h) and the Young's modulus (E) of the flexible plate, ISD4/(hE),is equal to or less than 10{circumflex over ( )}6 um3/GPa, e.g., lessthan 10{circumflex over ( )}5 um3/GPa, less then 10{circumflex over( )}4 um3/GPa or less than 10{circumflex over ( )}3 um3/GPa.

In some embodiments, one or both plates comprises a location markereither on a surface of or inside the plate, that provide information ofa location of the plate, e.g., a location that is going to be analyzedor a location onto which the blood should be deposited. In some cases,one or both plates may comprise a scale marker, either on a surface ofor inside the plate, that provides information of a lateral dimension ofa structure of the blood sample and/or the plate. In some embodiments,one or both plates comprises an imaging marker, either on surface of orinside the plate that assists an imaging of the sample. For example, theimaging marker could help focus the imaging device or direct the imagingdevice to a location on the device. In some embodiments, the spacers canfunction as a location marker, a scale marker, an imaging marker, or anycombination of thereof.

In some embodiments, on one of the sample surface, it further comprisesan enclosure-spacer that encloses a partial or entire VC samplesdeposited on the collection plate.

In some embodiments, the highly uniform thickness has a value equal toor less than 0.5 um. In some embodiments, the highly uniform thicknesshas a value in the range of 0.5 um to 1 um, 1 um to 2 um, 2 um to 10 um,10 um to 20 um or 20 um to 30 um.

In some embodiments, the thickness of the at least a part of VC sampleat the closed configuration is larger than the thickness of VC sampledeposited on the collection plate at an open configuration.

In some embodiments, the thickness of the at least a part of VC sampleat the closed configuration is less than the thickness of VC sampledeposited on the collection plate at an open configuration.

In some embodiments, wherein the spacers are fixed on a plate bydirectly embossing the plate or injection molding of the plate.

In some embodiments, wherein the materials of the plate and the spacersare selected from polystyrene, PMMA, PC, COC, COP, or another plastic.

In some embodiments, the inter-spacer spacing in the range of 1 um to 50um, 50 um to 100 um, 100 um to 200 um or 200 um to 1000 um.

In some embodiments, the VC sample is an exhaled breath condensate froma human or an animal.

In some embodiments, the spacers regulating the layer of uniformthickness have a filling factor of at least 1%, wherein the fillingfactor is the ratio of the spacer area in contact with the layer ofuniform thickness to the total plate area in contact with the layer ofuniform thickness.

In some embodiments, for spacers regulating the layer of uniformthickness, the Young's modulus of the spacers times the filling factorof the spacers is equal or larger than 10 MPa, wherein the fillingfactor is the ratio of the spacer area in contact with the layer ofuniform thickness to the total plate area in contact with the layer ofuniform thickness.

In some embodiments, for a flexible plate, the thickness of the flexibleplate times the Young's modulus of the flexible plate is in the range 60to 750 GPa-um.

In some embodiments, for a flexible plate, the fourth power of theinter-spacer-distance (ISD) divided by the thickness of the flexibleplate (h) and the Young's modulus (E) of the flexible plate, ISD4/(hE),is equal to or less than 10{circumflex over ( )}6 um3/GPa,

In some embodiments, one or both plates comprises a location marker,either on a surface of or inside the plate, that provide information ofa location of the plate.

In some embodiments, one or both plates comprises a scale marker, eitheron a surface of or inside the plate, that provide information of alateral dimension of a structure of the sample and/or the plate.

In some embodiments, one or both plates comprises an imaging marker,either on surface of or inside the plate, that assists an imaging of thesample.

In some embodiments, the spacers functions as a location marker, a scalemarker, an imaging marker, or any combination of thereof.

In some embodiments, the average thickness of the layer of uniformthickness is about equal to a minimum dimension of an analyte in thesample.

In some embodiments, the inter-spacer distance is 1 μm or less, 5 μm orless, 7 μm or less, 10 μm or less, 20 μm or less, 30 μm or less, 40 μmor less, 50 μm or less, 60 μm or less, 70 μm or less, 80 μm or less, 90μm or less, 100 μm or less, 200 μm or less, 300 μm or less, 400 μm orless, or in a range between any two of the values.

In some embodiments, the inter-spacer distance is substantiallyperiodic.

In some embodiments, the inter-spacer distance is aperiodic.

In some embodiments, the spacers are pillars with a cross-sectionalshape selected from round, polygonal, circular, square, rectangular,oval, elliptical, or any combination of the same.

In some embodiments, the spacers have are pillar shape and have asubstantially flat top surface, wherein, for each spacer, the ratio ofthe lateral dimension of the spacer to its height is at least 1.

In some embodiments, each spacer has the ratio of the lateral dimensionof the spacer to its height is at least 1.

In some embodiments, the minimum lateral dimension of spacer is lessthan or substantially equal to the minimum dimension of an analyte inthe sample.

In some embodiments, the minimum lateral dimension of spacer is in therange of 0.5 um to 100 um.

In some embodiments, the minimum lateral dimension of spacer is in therange of 0.5 um to 10 um.

In some embodiments, the spacers have a density of at least 100/mm2. Insome embodiments, the spacers have a density of at least 1000/mm2. Insome embodiments, at least one of the plates is transparent.

In some embodiments, at least one of the plates is made from a flexiblepolymer.

In some embodiments, for a pressure that compresses the plates, thespacers are not compressible and/or, independently, only one of theplates is flexible.

In some embodiments, the flexible plate for the device of any priorembodiment has a thickness in the range of 10 um to 200 um. In someembodiments, the plate thickness is 0.5 um, 1 um, 5 um, 10 um, 25 um, 50um, 75 um, 100 um, 125 um, 150 um, 175 um, 200 um, 250 um, or a rangebetween any of the two values.

In some embodiments, the variation in the thickness of the at least apart of the sample in a closed configuration of the plates is less than30%, 10%, 5%, 3% or 1%.

In some embodiments, the first and second plates are connected and areconfigured to be changed from the open configuration to the closedconfiguration by folding the plates.

In some embodiments, the first and second plates are connected by ahinge and are configured to be changed from the open configuration tothe closed configuration by folding the plates along the hinge.

In some embodiments, the first and second plates are connected by ahinge that is a separate material to the plates, and are configured tobe changed from the open configuration to the closed configuration byfolding the plates along the hinge

In some embodiments, the first and second plates are made in a singlepiece of material and are configured to be changed from the openconfiguration to the closed configuration by folding the plates.

In some embodiments, the layer of uniform thickness sample is uniformover a lateral area that is at least 100 um².

In some embodiments, the layer of uniform thickness sample is uniformover a lateral area that is at least 1 mm².

In some embodiments, the device is configured to analyze the sample in60 seconds or less.

In some embodiments, at the closed configuration, the final samplethickness device is configured to analyze the sample in 60 seconds orless.

In some embodiments, the device is configured to analyze the sample in180 seconds or less.

In some embodiments, at the closed configuration, the final samplethickness device is configured to analyze the sample in 180 seconds orless.

In some embodiments, the device further comprises, on one or both of theplates, one or a plurality of amplification sites that are each capableof amplifying a signal from the analyte or a label of the analyte whenthe analyte or label is within 500 nm from an amplification site.

In some embodiments, at the closed configuration, the final samplethickness device is configured to analyze the sample in 10 seconds orless.

In certain embodiments, the device of any prior embodiment include astep of amplifying the signal from the sample while being in a devicethat is in a closed configuration of the plates. The signalamplification is achieved without the use of by a physical processand/or biological/chemical amplification. Biological/chemicalamplification of the signal include enzymatic amplification of thesignal (e.g., used in enzyme-linked immunosorbent assays (ELISAs)) andpolymerase chain reaction (PCR) amplification of the signal. Thephysical signal amplification include plamonics, proximity dependentsurface amplification by metal structures or films, and others.

In some embodiments, the dry binding site comprises a capture agent.

In some embodiments, the dry binding site comprises an antibody ornucleic acid. In some embodiments, the releasable dry reagent is alabeled reagent. In some embodiments, the releasable dry reagent is afluorescently-labeled reagent. In some embodiments, the releasable dryreagent is a dye. In some embodiments, the releasable dry reagent is abeads. In some embodiments, the releasable dry reagent is a quantum dot.In some embodiments, the releasable dry reagent is afluorescently-labeled antibody.

In some embodiments, the first plate further comprises, on its surface,a first predetermined assay site and a second predetermined assay site,wherein the distance between the edges of the assay site issubstantially larger than the thickness of the uniform thickness layerwhen the plates are in the closed position, wherein at least a part ofthe uniform thickness layer is over the predetermined assay sites, andwherein the sample has one or a plurality of analytes that are capableof diffusing in the sample.

In some embodiments, the first plate has, on its surface, at least threeanalyte assay sites, and the distance between the edges of any twoneighboring assay sites is substantially larger than the thickness ofthe uniform thickness layer when the plates are in the closed position,wherein at least a part of the uniform thickness layer is over the assaysites, and wherein the sample has one or a plurality of analytes thatare capable of diffusing in the sample.

In some embodiments, the first plate has, on its surface, at least twoneighboring analyte assay sites that are not separated by a distancethat is substantially larger than the thickness of the uniform thicknesslayer when the plates are in the closed position, wherein at least apart of the uniform thickness layer is over the assay sites, and whereinthe sample has one or a plurality of analytes that are capable ofdiffusing in the sample.

In some embodiments, the releasable dry reagent is a cell stain. In someembodiments, the device further comprises a detector that is an opticaldetector for detecting an optical signal. In some embodiments, thedevice further comprises a detector that is an electrical detector fordetecting an electric signal.

In some embodiments, the device comprises discrete spacers that are notfixed to any of the plates, wherein at the closed configuration, thediscrete spacers are between the inner surfaces of the two plates, andthe thickness of the sample is confined by the inner surfaces of the twoplates, and regulated by the discrete spacers and the plates.

In some embodiments, the device further comprises a binding site thathas a chemical sensor that is made from a material selected from thegroup consisting of: silicon nanowire (Si NW); single-walled carbonnanotubes (SWCNT); random networks of carbon nanotubes (RN-CNTs);molecularly capped metal nanoparticles (MCNPs); metal oxidenanoparticles (MONPs); and chemically sensitive field-effect transistors(CHEM-FETs).

A system is provided herein for rapidly analyzing a vapor condensatesample using a mobile phone comprising:

-   -   (a) a device of any prior embodiments;    -   (b) a mobile communication device comprising:        -   i. one or a plurality of cameras for the detecting and/or            imaging the vapor condensate sample; and        -   ii. electronics, signal processors, hardware and software            for receiving and/or processing the detected signal and/or            the image of the vapor condensate sample and for remote            communication.

In some embodiments, the system further comprise a light source fromeither the mobile communication device or an external source.

In some embodiments, one of the plates has a binding site that binds ananalyte, wherein at least part of the uniform sample thickness layer isover the binding site, and is substantially less than the averagelateral linear dimension of the binding site.

In some embodiments, the system further comprising:

(d) a housing configured to hold the sample and to be mounted to themobile communication device.

In some embodiments, the housing comprises optics for facilitating theimaging and/or signal processing of the sample by the mobilecommunication device, and a mount configured to hold the optics on themobile communication device.

In some embodiments, an element of the optics in the housing is movablerelative to the housing.

In some embodiments, the mobile communication device is configured tocommunicate test results to a medical professional, a medical facilityor an insurance company.

In some embodiments, the mobile communication device is furtherconfigured to communicate information on the test and the subject withthe medical professional, medical facility or insurance company.

In some embodiments, the mobile communication device is furtherconfigured to communicate information of the test to a cloud network,and the cloud network process the information to refine the testresults.

In some embodiments, the mobile communication device is furtherconfigured to communicate information of the test and the subject to acloud network, the cloud network process the information to refine thetest results, and the refined test results will send back the subject.

In some embodiments, the mobile communication device is configured toreceive a prescription, diagnosis or a recommendation from a medicalprofessional.

In some embodiments, the mobile communication device is configured withhardware and software to:

-   -   a. capture an image of the sample;    -   b. analyze a test location and a control location in in image;        and    -   c. compare a value obtained from analysis of the test location        to a threshold value that characterizes the rapid diagnostic        test.

In some embodiments, at least one of the plates comprises a storage sitein which assay reagents are stored. In some embodiments, at least one ofthe cameras reads a signal from the CROF device. In some embodiments,the mobile communication device communicates with the remote locationvia a wifi or cellular network.

In some embodiments, the mobile communication device is a mobile phone.

A method is provided herein for rapidly analyzing an analyte in a sampleusing a mobile phone, comprising:

-   -   a) depositing a sample on the device of any prior embodiment;    -   b) assaying an analyte in the sample deposited on the device to        generate a result; and    -   c) communicating the result from the mobile communication device        to a location remote from the mobile communication device.

In some embodiments, the analyte comprises a molecule (e.g., a protein,peptides, DNA, RNA, nucleic acid, or other molecule), cells, tissues,viruses, and nanoparticles with different shapes.

In some embodiments, the analyte comprises white blood cell, red bloodcell and platelets. In some embodiments, the method comprises:

-   -   a. analyzing the results at the remote location to provide an        analyzed result; and    -   b. communicating the analyzed result from the remote location to        the mobile communication device.

In some embodiments, the analysis is done by a medical professional at aremote location. In some embodiments, the mobile communication devicereceives a prescription, diagnosis or a recommendation from a medicalprofessional at a remote location.

In some embodiments, the thickness of the at least a part of VC sampleat the closed configuration is larger than the thickness of VC sampledeposited on the collection plate at an open configuration.

In some embodiments, the thickness of the at least a part of VC sampleat the closed configuration is less than the thickness of VC sampledeposited on the collection plate at an open configuration.

In some embodiments, the assaying step comprises detecting an analyte inthe sample.

In some embodiments, the analyte is a biomarker. In some embodiments,the analyte is a protein, nucleic acid, cell, or metabolite. In someembodiments, the assay done in step (b) is a binding assay or abiochemical assay.

A method is provided herein for analyzing an analyte in a vaporcondensate sample comprising:

obtaining a device of any prior device embodiment;

depositing the vapor condensate sample onto one or both pates of thedevice;

placing the plates in a closed configuration and applying an externalforce over at least part of the plates; and

analyzing the analyts in the layer of uniform thickness while the platesare the closed configuration.

In some embodiments, the method comprises:

(a) obtaining a sample;

(b) obtaining a first and second plates that are movable relative toeach other into different configurations, wherein each plate has asample contact surface that is substantially planar, one or both platesare flexible, and one or both of the plates comprise spacers that arefixed with a respective sample contacting surface, and wherein thespacers have:

-   -   i. a predetermined substantially uniform height,    -   ii. a shape of pillar with substantially uniform cross-section        and a flat top surface;    -   iii. a ratio of the width to the height equal or larger than        one;    -   iv. a predetermined constant inter-spacer distance that is in        the range of 10 um to 200 um;    -   v. a filling factor of equal to 1% or larger; and

(c) depositing the sample on one or both of the plates when the platesare configured in an open configuration, wherein the open configurationis a configuration in which the two plates are either partially orcompletely separated apart and the spacing between the plates is notregulated by the spacers;

(d) after (c), using the two plates to compress at least part of thesample into a layer of substantially uniform thickness that is confinedby the sample contact surfaces of the plates, wherein the uniformthickness of the layer is regulated by the spacers and the plates, andhas an average value equal to or less than 30 um with a variation ofless than 10%, wherein the compressing comprises:

-   -   bringing the two plates together; and    -   conformable pressing, either in parallel or sequentially, an        area of at least one of the plates to press the plates together        to a closed configuration, wherein the conformable pressing        generates a substantially uniform pressure on the plates over        the at least part of the sample, and the pressing spreads the at        least part of the sample laterally between the sample contact        surfaces of the plates, and wherein the closed configuration is        a configuration in which the spacing between the plates in the        layer of uniform thickness region is regulated by the spacers;        and

(e) analyzing the in the layer of uniform thickness while the plates arethe closed configuration;

wherein the filling factor is the ratio of the spacer contact area tothe total plate area;

wherein a conformable pressing is a method that makes the pressureapplied over an area is substantially constant regardless the shapevariation of the outer surfaces of the plates; and

wherein the parallel pressing applies the pressures on the intended areaat the same time, and a sequential pressing applies the pressure on apart of the intended area and gradually move to other area.

In some embodiments, the method comprises removing the external forceafter the plates are in the closed configuration; and imaging theanalytes in the layer of uniform thickness while the plates are theclosed configuration; and counting a number of analytes or the labels inan area of the image.

In some embodiments, the method comprises removing the external forceafter the plates are in the closed configuration; and measuring opticalsignal in the layer of uniform thickness while the plates are the closedconfiguration.

In some embodiments, the inter-spacer distance is in the range of 20 umto 200 um. In some embodiments, the inter-spacer distance is in therange of 5 um to 20 um.

In some embodiments, a product of the filling factor and the Young'smodulus of the spacer is 2 MPa or larger.

In some embodiments, the surface variation is less than 50 nm.

In some embodiments, the method further comprising a step of calculatingthe concentration of an analyte in the relevant volume of sample,wherein the calculation is based on the relevant sample volume definedby the predetermined area of the storage site, the uniform samplethickness at the closed configuration, and the amount of target entitydetected.

In some embodiments, the analyzing step comprise counting the analyte inthe sample. In some embodiments, the imaging and counting is done by:

i. illuminating the cells in the layer of uniform thickness;

ii. taking one or more images of the cells using a CCD or CMOS sensor;

iii. identifying cells in the image using a computer; and

iv. counting a number of cells in an area of the image.

In some embodiments, the external force is provided by human hand. Insome embodiments, the method future comprises a dry reagent coated onone or both plates.

In some embodiments, the layer of uniform thickness sample has athickness uniformity of up to +/−5%.

In some embodiments, the spacers are pillars with a cross-sectionalshape selected from round, polygonal, circular, square, rectangular,oval, elliptical, or any combination of the same.

In some embodiments, the spacing between the spacers is approximatelythe minimum dimension of an analyte.

The method of any prior embodiment, wherein one or both plate samplecontact surfaces comprises one or a plurality of amplification sitesthat are each capable of amplifying a signal from the analyte or a labelof the analyte when the analyte or label is within 500 nm from anamplification site.

In some embodiments, the sample is exhale breath condensate. In someembodiments, the sample is a vapor from a biological sample, anenvironmental sample, a chemical sample, or clinical sample. In someembodiments, the analyte comprises a molecule (e.g., a protein,peptides, DNA, RNA, nucleic acid, or other molecules), cells, tissues,viruses, and nanoparticles with different shapes In some embodiments,the analyte comprises volatile organic compounds (VOCs). In someembodiments, the analyte comprises nitrogen, oxygen, CO2, H2O, and inertgases. In some embodiments, the analyte is stained.

In some embodiments, on one of the sample surface, it further comprisesan enclosure-spacer that encloses a partial or entire VC samplesdeposited on the collection plate. In some embodiments, the highlyuniform thickness has a value equal to or less than 0.5 um. In someembodiments, the highly uniform thickness has a value in the range of0.5 um to 1 um. In some embodiments, the highly uniform thickness has avalue in the range of 1 um to 2 um. In some embodiments, the highlyuniform thickness has a value in the range of 2 um to 10 um. In someembodiments, the highly uniform thickness has a value in the range of 10um to 20 um. In some embodiments, the highly uniform thickness has avalue in the range of 20 um to 30 um.

In some embodiments, on one of the sample surface, it further comprisesan enclosure-spacer that encloses a partial or entire VC samplesdeposited on the collection plate.

Embodiment (EBC)-1. SiEBCA (Single-Drop Exhaled Breath CondensateCollector and Analyzer)

Exhaled breath condensate (EBC) analysis is a noninvasive method ofdetecting biomarkers, mainly coming from the lower respiratory tract.EBC is collected during quiet breathing, as a product of cooling andcondensation of the exhaled aerosol.

An exemplary method of collecting exhaled breath condensate (EBC) usinga SiEBCA (Single-drop EBC Collector/Analyzer), as illustrated in FIG. 1, comprises the basic steps:

(1) exhaling breath onto the collection plate (FIG. 1-1 ). A subject(e.g. human) breathe onto a plate, termed “collection plate”, and thebreath condenses into EBC, which are in droplets with different sizes,depending on the breathing time. For a short breathing time mostdroplets are separated from each other. The surface of the collectionplate that collects the EBC is termed the sample surface;

(2) placing a cover plate over the collection plate and pressing themtogether (FIG. 1-2 ). A cover plate with spacers (which are used forregulating the spacing between the cover plate and the substrate plate)is placed on top of the sample surface; and

(3) pressing plates into a “Closed-Configuration (FIG. 1-3 ). The coverplate and the substrate are compressed together with at least a part ofthe EBC between the plates.

In the method of FIG. 1 , the initial droplets are pressed into a thinlayer EBC of a thickness that is regulated by the plates and spacers(not shown).

One reason for using the wording of “single drop” in SiEBCA is that inprinciple, the SiEBCA can detect and analyze a single drop of EBCdeposited on the plate.

In the description of the present invention, “the substrate plate” and“the cover plate” are respectively interchangeable with “the firstplate” and “the second plate”.

In some embodiments, the plates are cooled to reduce the evaporation ofcollected EBC.

A1 A method of collecting EBC, as a basic embodiment of the presentinvention for collecting EBC from a subject, as illustrated in FIG. 1 ,comprises the steps:

(a) obtaining a collection plate and a cover plate that are movablerelative to each other into different configurations, wherein one orboth of the plates comprise spacers (not shown in FIG. 1 but in FIG. 2 )that are fixed with the respective plate, and have a predeterminedaverage height of 100 um or less;

(b) depositing, when the plates are configured in an open configuration,an EBC sample by exhaling breath from a subject toward the collectionplate, wherein:

-   -   (i) the exhaled breath condensates on a collection surface of        the collection plate to form droplets and/or puddles that have        different lateral sizes and different heights, depending upon        the surface wetting properties of the collection surface; and    -   (ii) the open configuration is a configuration in which the two        plates are either partially or completely separated apart and        the spacing between the plates is not regulated by the spacers;        and

(c) after (b), bringing the cover plate over the collection surface ofthe collection plate and then bringing the two plates into a closedconfiguration by pressing the plates, wherein:

-   -   (i) the closed configuration is a configuration, in which: at        least a part of the spacers are between the cover plate and the        collection plate, and a relevant area of the collection surface        of the collection plate is covered by the cove plate and has a        plate spacing that is regulated by the spacers; and    -   (ii) at the closed configuration, in the relevant area,        substantial number or all of the droplets or puddles formed in        step (b) at the open configuration merge into puddle(s) that (1)        have much larger lateral size but in a smaller number than the        open configuration and (2) touch both inner surfaces of the        cover plate and the collection plate, thereby the thickness of        the puddle(s) is confined by the inner surfaces of the plates        and equal to the spacing between the inner surfaces, and the        total surface area of the deposited EBC exposed to the ambient        is significantly reduced;    -   wherein the plate spacing is the spacing between the two inner        surfaces (the two surfaces facing each other) of the cover plate        and the collection plate, the relevant area is a portion or        entire surface of the collection surface, and the collection        surface is a portion or entire surface of the collection plate.

From our experiments (described in details in Examples), we found thatthe final form of the EBC collected by SiEBCA when the plates are in theclosed configuration depends upon the spacer height. Experimentally, wefound, as illustrated in FIG. 2 , that:

(1) At the closed configuration of the SiEBCA, if the spacing betweenthe inner surfaces of the plates is less than the average height of theEBC droplets or puddles at the open configuration, the EBC droplets orpuddles are compressed by the collection plate and the cover plate intoa continuous film of a thickness thinner than at the open configuration,and also air pockets may exist in the film; and

(2) otherwise (i.e. if the spacing is equal to or larger than that theaverage height at the open configuration) the droplets and/or puddlesself-emerged into discrete puddles that are fewer in number but largerin lateral size (area) than that in the open configuration, touch bothsample contact (inner) surfaces of the cover plate and the collectionplate, and have the thickness confined by the inner surfaces of theplates and equal to the spacing between the inner surfaces. In thiscase, the EBC sample thickness at the closed configuration is equal toor larger than the EBC sample average thickness at the openconfiguration. The increase in the EBC puddle thickness at the closedconfiguration, as we observed experimentally, are due to theinteractions between the plates and the EBC sample.

EBC-1.2. Device for EBC Collection

FIG. 4 . is an illustration of an embodiment of the devices and themethods of a SiEBCA (Single-drop EBC Collector/Analyzer): (a) having afirst plate and a second plate, wherein one or both plate has spacers(shown here: only the first plate has spacers); (b) depositing a sample(only one of many EBC droplets is shown) on the first plate (shown), orthe second plate (not shown), or both (not shown) at an openconfiguration; and (c) (i) using the two plates to spread the sample(the sample flow between the plates) and reduce the sample thickness,and (ii) using the spacers and the plate to regulate the samplethickness at the closed configuration.

A2 A device of collecting EBC, as a basic embodiment of the presentinvention for collecting EBC sample from a subject, as illustrated inFIG. 1 , comprises:

-   -   i. a first plate and a second plate, wherein the plates are        movable relative to each other into different configurations,        and one or both plates are flexible;    -   ii. a sample contact area on the respective surface of each of        the plates for contacting EBC sample,    -   iii. spacers on one or both of the plates, wherein the spacers        are fixed with a respective plate, have a predetermined        substantially uniform height of 30 um or less and a        predetermined constant inter-spacer distance that is 250 um or        less, and wherein at least one of the spacers is inside the        sample contact area;    -   wherein one of the configurations is an open configuration, in        which: the two plates are separated apart, the spacing between        the plates is not regulated by the spacers, and the EBC sample        is deposited on one or both of the plates from a subject; and    -   wherein another of the configurations is a closed configuration        which is configured after the EBC sample deposition in the open        configuration; and in the closed configuration: at least part of        the EBC sample is compressed by the two plates into a layer of        highly uniform thickness, wherein the uniform thickness of the        layer is in contact with and confined by the inner surfaces of        the two plates and is regulated by the plates and the spacers.

In some embodiments of paragraphs A1 and A2, the deposition of the EBCsample is by directly exhaling from a subject to one of the plates.

In some embodiments of paragraphs A1 and A2, the deposition of the EBCsample is by directly exhaling from a subject to both of the plates.

“Covering time delay” means a time period that it takes from the step(b) EBC deposition of paragraph A1 to the end of the step (c) ofparagraph A1 that brings the cover plate and the collection plate to aclosed configuration.

In the method of paragraph A1, the covering time delay should be asshort as possible to reduce the evaporation of deposited EBC. In onepreferred embodiment, the covering time delay is equal to or less than 2sec. In another preferred embodiment, the covering time delay is equalto or less than 5 sec. In another preferred embodiment, the coveringtime delay is equal to or less than 10 sec. In another preferredembodiment, the covering time delay is equal to or less than 30 sec. AndIn another preferred embodiment, the covering time delay is in the rangeof 30 sec to 300 sec (e.g. 30 to 60 sec, 60 sec to 120 sec, or 120 secto 300 sec).

EBC-1.3. Significant reduction of EBC evaporation rate. One keyadvantage of the method and device of paragraph A1 and A2 is that,compared to the open configuration of the collection plate and the coverplate, the closed configuration of the plates significantly reduces thesurface area of the EBC exposed to the ambient, and hence significantlyreduces the EBC sample evaporation rate and significantly increases thetime that EBC sample is in liquid form (i.e. the time that EBC sample isnot completely evaporated). For example, we have observed that thedrying time (the time it takes for the EBC sample to dry out completely)increased from 30 secs at an open configuration to 70 mins, a factor of140 times longer.EBC-1.4. Guard ring (Enclosed Spacers). To further reduce the EBC sampleevaporation rate, the enclosed spacers or the guard rings can be used tosurround the sample to seal off the sample from the ambient. The guardring can circle an area that is the same as, or larger or smaller thanthe EBC sample deposited at the open configuration. The guard ring canbe configured to further divide an EBC sample into multiple chambers(FIG. 5 ).

FIG. 5 is an illustration of a SiEBCA with both “open spacer” andenclosed spacer, where the open spacer is a post (pillar) while theenclosed spacer is a ring. The enclosed spacer reduces the evaporationof the EBC collected inside the enclosed spacer, since at the closedconfiguration, the enclosed spacer, the cover plate and the collectionplate form an enclosed chamber. If there are only the open spacers butnot an enclosed spacer, at the plate closed configuration, the collectedEBC still evaporates from the edge of the film formed by the EBC,although such evaporation is much slower than that without a coverplate.

In the method and the device of paragraph A1 and A2, the spacers can bean open spacer(s), an enclosed spacer(s), or a combination of thereof.

Details of the devices and methods to keep the EBC thickness uniform aregiven in the other part of the disclosure.

Embodiment EBC-2. EBC Analysis

A-3 A method of analyzing EBC from a subject for analyzing EBC of asubject comprising:(a) obtaining a device of any prior embodiments;(b) depositing, when the plates are configured in an open configuration,an EBC sample by exhaling breath from a subject toward the collectionplate, wherein:

-   -   (i) the exhaled breath condensates on a collection surface of        the collection plate to form droplets and/or puddles that have        different lateral sizes and different heights, depending upon        the surface wetting properties of the collection surface; and    -   (ii) the open configuration is a configuration in which the two        plates are either partially or completely separated apart and        the spacing between the plates is not regulated by the spacers;        (c) after (b), bringing the cover plate over the collection        surface of the collection plate and then bringing the two plates        into a closed configuration by pressing the plates, wherein:    -   (i) the closed configuration is a configuration, in which: at        least a part of the spacers are between the cover plate and the        collection plate, and a relevant area of the collection surface        of the collection plate is covered by the cove plate and has a        plate spacing that is regulated by the spacers; and    -   (ii) at the closed configuration, in the relevant area,        substantial number or all of the droplets or puddles formed in        step (b) at the open configuration merge into puddle(s) that (1)        have much larger lateral size but in a smaller number than the        open configuration and (2) touch both inner surfaces of the        cover plate and the collection plate, thereby the thickness of        the puddle(s) is confined by the inner surfaces of the plates        and equal to the spacing between the inner surfaces, and the        total surface area of the deposited EBC exposed to the ambient        is significantly reduced; and        (d) analyzing the EBC,

wherein the plate spacing is the spacing between the two inner surfaces(the two surfaces face each other) of the cover plate and the collectionplate, the relevant area is a portion or entire surface of thecollection surface, and the collection surface is a portion or entiresurface of the collection plate.

The collection plate generally is held at a temperature the same as theambient, but in some embodiments, the temperatures can be different fromthe ambient, either higher or lower depending upon the goal of thecollection. For example, a temperature lower than the ambient may beused for reducing the EBC evaporation; and a temperature higher than theambient many bused for evaporating more than that at the ambienttemperature is needed.

The present invention provides that EBC samples and the target analytestherein can be analyzed by a variety of analysis techniques, dependingon the sample volume, species and abundance of the target analyte(s). Anon-exhaustive and mutually non-exclusive list of these analysistechniques include spectrometry (e.g., mass spectrometry (MS), ionmobility spectrometry, proton transfer spectrometry, optical absorptionspectroscopy systems, and spectrofluorimetry), enzyme based techniques,immunoassay methods (e.g., ELISA, radioimmunoassay, and immune sensors),electronic noses, 2-dimensional protein gel electrophoresis (2D-PEG)followed by chromatographic proteomic microanalysis, nucleic acid(including amplification by PRC (polymerase chain reaction) andisothermal amplification of nucleic acid) by and various types ofchemical sensors.

4. Additional Exemplary Analysis by SiVC with or without Spacers

Another significant advantage of the present invention is that themethod and the device of paragraph A1-2, in some embodiments, are usedas an EBC analyzer by itself directly or with certain modifications. TheEBC analyzer analyzes one or a plurality of target analytes in the EBC.The target analytes are further discussed in Section 3.

In some embodiments, the modifications made to the method and device ofparagraphs A1-A2 include, but not limited to, the following, which canused alone (individually) or in combinations:

(1) Binding Sites. One or both of the plates have one or plurality ofbinding site Each (type) of the reagents are in either in well separatedlocations (the well separation will be defined later).

(2) Storage sites. One or both of the plates have one or plurality ofbinding site Each (type) of the regents are in either in well separatedlocations (the well separation will be defined later).

(3) Amplification site.

(4) Muiltplexing of analyte detections.

More details of the binding sites, storage sites, amplification sites,and multiplexing sites, as well as their usage for VC and EBC analysisare given in the other part of the disclosure.

In addition to the various embodiments detailed in other parts of thedisclosure, in some embodiments, the modifications made to the methodand the device of paragraphs A1-A2 included the use of chemical sensorsas binding agents in the binding site for sensing target analytes in theEBC sample, in particular VOC compounds. These chemical sensors include,but not limited to. sensors based on conducting polymers and metaloxides, surface acoustic wave sensors, and optical sensors. In someembodiments, they involve the use of nanotechnology (gold nanoparticles,nanowires, and nanotubes) for the design of VOC sensors. In someembodiments, the chemical sensors give output in the forms of colorchange, fluorescence, conductivity, vibration, sound, or any combinationthereof, following the interaction of the VOC compound with the sensor.

In particular embodiments, the chemical sensor is made from materials,including, but not limited to, silicon nanowire (Si NW); single-walledcarbon nanotubes (SWCNT); random networks of carbon nanotubes (RN-CNTs);molecularly capped metal nanoparticles (MCNPs); metal oxidenanoparticles (MONPs); and chemically sensitive field-effect transistors(CHEM-FETs) that are respectively functionalized for the detection ofVOCs or other analytes in the EBC sample. The term “functionalization”(or “functionalized”) as used herein refer to the modification to (orthe modified properties of) a material that enables it to have specificinteraction (e.g., binding, chemical reaction) with certain type(s) oftarget analytes and bring about a signal indicative of such interaction,thereby to sense the existence and/or abundance of the target analyte.The term “functional group” as used herein refers to the chemical groupthat is chemically or physically coupled to the material thatfunctionalizes the material.

In some embodiments, the functional group is made from a protein, anamino acid, a nucleic acid, a lipid, a carbohydrate, a metabolite, anyother organic compound, or any other inorganic compound that is able tointeract with target analyte in the EBC sample.

In the art of EBC detection, in particular the sampling/sensing of theVOC compound, it is challenging to achieve high selectivity againstindividual components of the sample. Many available reagents (sensors,e.g., a particular type of chemical sensor or functional group, or aparticular antibody) are not particularly sensitive to one type oftarget analytes, rather selective to a certain range of target analytes.A solution to this is to utilize an array of different cross-reactivesensors in couple with data processing system capable of patternrecognition to virtually separate and identify individual componentsbased on the output pattern/parameters of the sensors. The term“cross-reactive” as used herein refers to the fact that a number ofreagents/sensors are selectively responsive to a respective range ofanalytes and the responsive range of each reagent/sensor overlapspartially with at least one of the other reagents/sensors.

In some embodiments, the device and method of paragraphs A1-A2 include avariety of reagents (sensors), many of which are cross-reactive, and thedevice and method are utilized in conjunction with a data processingsystem, including both hardware and software that are capable of patternrecognition. In particular, the data processing methods embodied by thedata processing system include, but not limited to, principal componentanalysis, cluster analysis, discriminant function analysis, geneticalgorithms, neutral network algorithms, and any other machine learningalgorithms.

In some other embodiments, the method and device of prior embodimentsare used for collecting EBC samples for analysis by other EBCanalyzer(s) according to any of the aforementioned analysis techniquesor any combination thereof.

Another advantage of the method and device of prior embodiments a isthat the volume of the collected sample is ready to be quantified, or itis easy to quantitatively collect a pre-determined volume of sample forfurther analysis. As detailed in other part of the disclosure, in someembodiments, a continuous film of the sample is formed after bringingthe plates in to the closed configuration. The relevant volume of sampleis therefore readily and precisely determined by timing the relevantarea by the thickness of the sample film, which is equal to the platespacing. In some embodiments, the plate spacing is equal to or deviatingwith a small variation from the spacer height. In some embodiments, thespacers have a uniform height, and the sample film has a uniformthickness equal to the uniform height.

In some embodiments, the analyzing step of paragraph A0 to A3comprises: 1) at the closed configuration, taking a first volume of thesample having a first lateral area out of the sample collected inbetween the cover plate and the collection plate; 2) analyzing the firstvolume of the sample by an analyzer. As discussed above, the firstvolume of the sample is equal to the product of the first lateral areaand the plate spacing. In some embodiments, the first lateral area ispre-determined or pre-known. For instance, the device features visualmarks that define or border a certain area on the cover plate, thecollection plate, or both, within the overlapping area between the twoplates, and the first volume of sample is taken according to the marks,e.g., by cutting out along the marks. In this case, the sample needs tobe a continuous film with no or little empty space within the boundaryset by the visual marks. In some embodiments, the first lateral area ismeasured during or after the taking step.

In some other embodiments, the EBC samples collected according to themethod and device of paragraphs A1-A2 are analyzed by a combination ofboth the EBC device itself and other analyzers.

Embodiment EBC-3. Exemplary Applications of SiVC/SiEBC

EBC-3.1. Analysis of EBC

Breath tests are among the least invasive methods available for clinicaldiagnosis, disease state monitoring, health monitoring and environmentalexposure assessment.

EBC analysis can be used for detection of inflammatory markers, whichreflect the state of chronic airways diseases such as chronicobstructive pulmonary disease (COPD), asthma, and cystic fibrosis (CF).EBC analysis can also be used for identification of metabolic,proteomic, and genomic fingerprints of breathing, aiming for an earlydiagnosis of not only respiratory, but also systemic diseases.

A breath matrix from a subject is a mixture of nitrogen, oxygen, CO2,H2O, and inert gases. The remaining small fraction consists of more than1000 trace volatile organic compounds (VOCs) with concentrations in therange of parts per million (ppm) to parts per trillion (ppt) by volume.In terms of their origin, these volatile substances may be generated inthe body (endogenous) or may be absorbed as contaminants from theenvironment (exogenous). The composition of VOCs in breath varies widelyfrom person to person, both qualitatively and quantitatively.

Although the number of VOCs found to date in human breath is more than1000, only a few VOCs are common to all humans. These common VOCs, whichinclude isoprene, acetone, ethane, and methanol, are products of coremetabolic processes and are very informative for clinical diagnostics.The bulk matrix and trace VOCs in breath exchange between the blood andalveolar air at the blood-gas interface in the lung. One exception isNO, which is released into the airway in the case of airwayinflammation.

The endogenous compounds found in human breath, such as inorganic gases(e.g., NO and CO), VOCs (e.g., isoprene, ethane, pentane, acetone), andother typically nonvolatile substances such as isoprostanes,peroxynitrite, or cytokines, can be measured in breath condensate.Testing for endogenous compounds can provide valuable informationconcerning a possible disease state. Furthermore, exogenous molecules,particularly halogenated organic compounds, can indicate recent exposureto drugs or environmental pollutants.

Volatile Organic Compounds (VOCs) are organic substances that have ahigh vapor pressure and therefore evaporate at room temperature. TheVOCs that may be assayed as target analytes by the methods and devicesprovided by the present invention include, but not limited to,biologically generated VOCs (e.g., terpenes, isoprene, methane, greenleaf volatiles) and anthropogenic VOCs (e.g., typical solvents used inpaints and coatings, like ethyl acetate, glycol ethers, and acetone,vapors from adhesives, paints, adhesive removers, building materials,etc., like methylene chloride, MTBE, and formaldehyde, chlorofurocarbonsand perchloroethylene used in dry cleaning, vapor and exhaustive gasfrom fossil fuels, like benzene and carbon monoxide).

Detailed discussion on certain breath markers for diseases and otherhealth conditions is given in Table 1.

Besides the diseases listed in Table 1, various VOCs contained inexhaled breath have also been linked to different types of cancers. Anon-exclusive list of breath VOCs identified as biomarkers for cancersis shown in Table 2.

Besides some of the non-volatile compounds listed in Table 1, variousnon-volatile compounds have also been lined to or identified asbiomarkers of various diseases/conditions. Among these, a particularapplication of the device and method provided by the present disclosureis to assay the glucose level in EBC. Other applications include, butnot limited to, detecting the levels of nitrogen reactive species,arachidonic acid metabolites (e.g., isoprostanes, leukotrienes,prostanoids), cytokines, glutathione, proteins and metabolites, smallmolecules (e.g., chloride, sodium, potassium, urea, and small organicacids), and pH.

In some embodiments, the devices and methods of the present inventionalso find applications in the detection of drugs of abuse in EBC sample.The drugs of abuse to be detected using the devices and methods of thepresent invention include, but not limited to, ethanol, cannabis,methadone, amphetamine, methamphetamine,3,4-methylenedioxymethamphetamine, codeine, 6-acetylmorphine, diazepam,oxazepam, morphine, benzoylecgonine, cocaine, buprenorphine andtetrahydrocannabinol.

TABLE 1 Breath markers in certain diseases or conditionsDisease/Condition Breath Marker Diabetes/diabetic ketoacidosis Acetone,Ethylbenzene, Xylene, Touluene, Ethane, Pentane, Propane, Isoprene,Ethanol, Methanol, Isopropanol, 2,3,4-Trimethylhexane, 2,6,8-Trimethyldecane, Tridecane, Undecane Helicobacter pylori infectionAmmonia, volatile organic compounds Uremia/kidney failure Dimethylamine,trimethylamine Liver disease Dimethylamine, trimethylamine Liver diseaseEthanethiol, dimethylsulfide, hydrogen disulfide Liver cirrhosisAcetone, styrene, dimethylsulfide, dimethylselene Liver disease/fetorhepaticus C2-C5 Aliphatic acids, methylmercaptan Angina, ischemic heartdisease Alkanes, methylated alkanes Heart-transplant rejectionMethylated alkane contour Rheumatoid arthritis Pentane Allograftrejection CS2 Oxidative stress NO, CO, nitrosothiol, 8-isoprostane,4-hydroxy-2- nonenal, malondialdehyde, hydrogen peroxide Chronicobstructive pulmonary NO, CO, nitrosothiol, hydrogen peroxide diseaseRhinitis, rhinorrhea chronic cough NO Asthma Pentane, ethane,8-isoprostane, NO, pH, H2O2, leukotrienes (e.g., LTs, Cys-LTs, LTE4), 8-Isoprostane, PGE2, ILs, IL-4, IL-5, IL-6, IL-8, IL-10, IL-17, INF-▭,RANTES, MIP▭, MIP▭, TNF-▭, TGF-▭, ET-1, Cytokeratine 1, MDA, ADMA,CCL11, hs-CRP, sICAM-1 Cystic fibrosis NO, CS2, leukotrienes (e.g.,LTE4), pH, Nitrotyrosine, Nitrites, Nitric oxide, 8-Isoprostane, IL-6,IL-8, IL-5, TNF-▭ Bronchiectasis NO Lung cancer Alkanes, monomethylatedalkanes, nitric oxide Lung carcinoma Acetone, methylethylketone,n-propanol, alkanes, aniline, o-toluidine Breast cancer 2-propanol,2,3-dihydro-1-4(1H)-quinazolinone, 1- phenyl-ethanone, heptanal,isopropyl myristate Idiopathic pulmonary fibrosis (IPF) 8-Isoprostane,H2O2, 3-nitrotyrosine, NOx, docosatetraenoyl-LPA Pulmonary arterialhypertension Natriuretic peptide, pro-BNP, ET-1, 6-keto-PGF1α, (PAH)8-isoprostane, IL-6 Sarcoidosis 8-Isoprostane, Cys-LTs, Neopterin, TGF-▭Obstructive SleepApnea Syndrome (OSA) Pediatric patients 8-Isoprostane,IL-6, LTB4, Cys-LTs, H2O2, Uric salts Adult patients 8-Isoprostane,IL-6, TNF-▭, pH, H2O2, ICAM-1, IL-8 Systemic Lupus Erythematosus (SLE)IL-6, IL-8, IL-10 Chronic Renal Disease (CRD) pH, Nitrites, Nitrates,H2O2

TABLE 2 VOCs from exhaled breath that are identified in biomarkers ofvarious cancers Disease Compound name Lung cancer 1,3-Cyclopentadiene,1-methyl- 1-Cyclopentene 2,3-Butanedione 2-Butanol, 2,3-dimethyl-2-Butanone (methyl ethyl ketone) 2-Butanone, 3-hydroxy- 2-Butene,2-methyl- 3-Butyn-2-ol Acetophenone Benzaldehyde Benzene, cyclobutyl-Butane, 2-methyl- Butyl acetate Ethylenimine Isoquinoline,1,2,3,4-tetrahydro- Methyl propyl sulfide n-Pentanal n-UndecaneUndecane, 3,7-dimethyl- Urea, tetramethyl- Cyclopentane Acetone Methylethyl ketone n-Propanol 1,1′-(1-Butenylidene)bis benzene1-Methyl-4-(1-methylethyl)benzene 2,3,4-Trimethyl hexane 3,3-Dimethylpentane Dodecane 1,3-Butadiene, 2-methyl-(isoprene) 1-Heptene 1-HexeneBenzene Benzene, 1,2,4-trimethyl- Benzene, 1,4-dimethyl Benzene,1-methylethenyl- Benzene, propyl- Cyclohexane Cyclopentane, methyl-Cyclopropane, 1-methyl-2-pentyl- Decane Heptane, 2,2,4,6,6-pentamethylHeptane, 2,4-dimethyl Heptane, 2-methyl Hexanal Methane,trichlorofluoro- Nonane, 3-methyl- Octane, 3-methyl- styrene(ethenylbenzene) Undecane Butane Decane, 5-methyl Heptane Hexane,2-methyl Hexane, 3-methyl Octane, 4-methyl Pentane Tridecane, 3-methylTridecane, 7-methyl 1,1-Biphenyl, 2,2-diethyl- 1,2-Benzenedicarboxylicacid, diethyl ester 1,5,9-Cyclododecatriene, 1,5,9-trimethyl-10,11-Dihydro-5H-dibenz[b,f]azepine 1H-Indene,2,3-dihydro-1,1,3-trimethyl-3-phenyl- 1-Propanol 2,4-Hexadiene,2,5-dimethyl- 3-Pentanone, 2,4-dimethyl- 2,5-Cyclohexadiene-1,4-dione,2,6-bis(1,1-dimethylethyl)- Benzene, 1,1-oxybis- Benzoic acid,4-ethoxy-, ethyl ester Decane, 4-methyl- Furan, 2,5-dimethyl-Pentan-1,3-dioldiisobutyrate, 2,2,4-trimethyl Propanoic acid, 2-methyl-,1-(1,1-dimethylethyl)-2-methyl-1,3- propanediyl estertrans-Caryophyllene 1,2,4,5-Tetroxane, 3,3,6,6-tetraphenyl- 1H-Indene,2,3-dihydro-4-methyl- 1-Propene, 1-(methylthio)-, (E)-2,2,4-Trimethyl-1,3-pentanediol diisobutyrate2,2,7,7-Tetramethyltricyclo-[6.2.1.0(1,6)]undec-4-en-3-one2,3-Hexanedione 2,5-Cyclohexadien-1-one,2,6-bis(1,1-dimethylethyl)-4-ethylidene 2-Methyl-3-hexanone4-Penten-2-ol 5,5-Dimethyl-1,3-hexadiene5-Isopropenyl-2-methyl-7-oxabicyclo[4.1.0]heptan-2-ol9,10-Anthracenediol, 2-ethyl- Anthracene, 1,2,3,4-tetrahydro-9-propyl-Benzene, 1,1-(1,2-cyclobutanediyl)bis, cis- Benzene,1,1-[1-(ethylthio)propylidene]bis- Benzene, 1,1-ethylidenebis, 4-ethyl-Benzophenone Bicyclo[3.2.2]nonane-1,5-dicarboxylic acid, 5-ethyl esterCamphor Ethane, 1,1,2-trichloro-1,2,2-trifluoro- Furan,2-[(2-ethoxy-3,4-dimethyl-2-cyclohexen-1-ylidene)methyl]- Isomethylionone Isopropyl alcohol Pentanoic acid,2,2,4-trimethyl-3-carboxyisopropyl, isobutyl ester Propane,2-methoxy-2-methyl- α-Isomethyl ionone Butanal Heptanal Nonanal OctanalPentanal Propanal Ethylbenzene Octane Pentamethylheptane Toluene2-Methylpentane Isoprene Xylenes total Styrene Aniline o-Toluidine1-Butanol 3-Hydroxy-2-butanone 2,6,10-Trimethyltetradecane2,6,11-Trimethyldodecane 2,6-Dimethylnaphthalene 2,6-Di-tert-butyl-,4-methylphenol 2-Methylhendecanal 2-Methylnaphthalene 2-Pentadecanone3,7-Dimethylpentadecane 3,8-Dimethylhendecane 4-Methyltetradecane5-(1-Methyl)propylnonane 5-(2-Methyl)propylnonane 5-Butylnonane5-Propyltridecane 7-Methylhexadecane 8-Hexylpentadecane8-Methylheptadecane Eicosane Hexadecanal Nonadecane NonadecanolTridecane Tridecanone Formaldehyde (methanal) Isopropanol Breast cancer2,3,4-Trimethyldecane 2-Amino-5-isopropyl-8-methyl-1-azulenecarbonitrile3,3-Dimethyl pentane 5-(2-Methylpropyl)nonane 6-Ethyl-3-octyl ester2-trifluoromethyl benzoic acid Nonane Tridecane, 5-methyl Undecane,3-methyl Pentadecane, 6-methyl Propane, 2-methyl Nonadecane, 3-methylDodecane, 4-methyl Octane, 2-methyl 1-Phenylethanone2,3-Dihydro-1-phenyl-4(1H)-quinazolinone 2-Propanol Heptanal Isopropylmyristate (+)-Longifolene 1,3-Butadiene, 2-methyl- 1,4-Pentadiene1H-Cycloprop[e]azulene, decahydro-1,1,7-trimethyl-4-methylene-1-Octanol, 2-butyl- 2,5-Cyclohexadiene-1,4-dione,2,6-bis(1,1-dimethylethyl)- 2,5-Di-tert-butyl-1,4-benzoquinone2-Hexyl-1-octanol3-Ethoxy-1,1,1,5,5,5-hexamethyl-3-(trimethylsiloxy)trisiloxane Aceticacid, 2,6,6-trimethyl-3-methylene-7-(3-oxobutylidene)oxepan-2- yl esterBenzene, 1,2,3,5-tetramethyl- Benzene, 1,2,4,5-tetramethyl- Benzene,1-ethyl-3,5-dimethyl- Benzoic acid, 4-methyl-2-trimethylsilyloxy-,trimethylsilyl ester Cyclohexene, 1-methyl-5-(1-methylethenyl)-Cyclohexene, 1-methyl-5-(1-methylethenyl)-, (R)- Cyclopropane,ethylidene Cyclotetrasiloxane, octamethyl- D-Limonene Dodecane Dodecane,2,6,11-trimethyl- Dodecane, 2,7,10-trimethyl- Longifolene-(V4)Pentadecane Tetradecane Tridecane Trifluoroacetic acid, n-octadecylester Undecane Colon cancer 1,1′-(1-Butenylidene)bis benzene1,3-Dimethylbenzene 4-(4-Propylcyclohexyl)-4′-cyano[1,1′-biphenyl]-4-ylester benzoic acid 2-Amino-5-isopropyl-8-methyl-1-azulenecarbonitrile[(1,1-Dimethylethyl)thio]acetic acid Esophagogastric Ethylphenol cancerHexanoic acid Methylphenol Phenol Gastric cancer 2-ButoxyethanolIsoprene 2-Propenenitrile 6-Methyl-5-hepten-2-one Furfural(furfuraldehyde) Head and neck 4,6-Dimethyldodecane cancer5-Methyl-3-hexanone 2,2-Dimethyldecane Limonene2,2,3-Trimethyl-exobicyclo[2.2.1]heptane 2,2-Dimethyl-propanoic acidAmmonium acetate 3-Methylhexane 2,4-Dimethylheptane 4-Methyloctanep-Xylene 2,6,6-Trimethyloctane 3-Methylnonane Liver cancer3-Hydroxy-2-butanone Styrene Decane Ovarian cancer Decanal NonanalStyrene 2-Butanone Hexadecane Prostate cancer Toluene p-Xylene2-Amino-5-isopropyl-8-methyl-1-azulenecarbonitrile 2,2-Dimethyldecane

EBC-3.2. Collection and Analysis of Other Vapor Condensates.

Certain embodiments of the present invention are related to theapplications of the SiEBCA methods and devices for collection andanalysis of the vapor condensates other than the EBC. The othermoistures include, but not limited to, fog, clouds, steams, etc. Thetarget analysis of these vapor condensates can be for different purposeenvironmental monitoring, emission control, etc. In some embodiments,the sample is a vapor from a biological sample, an environmental sample,a chemical sample, or clinical sample.

EBC-3.3. Automatic and High Throughput.

In certain embodiments, the devices and methods of the present inventionare automatic and high speed, where the steps are performed by machines.In some embodiments, the plates are in the form of roll of sheets andare controlled by rollers to put certain area of the plates into an openconfiguration or a closed configuration.

EBC-3.4. Identification and Validation of Markers in Vapor Condensate

In certain embodiments, the devices and methods of the present inventionare particularly useful for the identification and validation ofbiomarkers for human diseases/conditions, or other markers forenvironmental, food safety, or other conditions/events. Due to the ease,fast speed, small sample volume, and multiplexing potential of thepresent devices and methods, it is easy to adapt the present device forhigh-throughput and even automatic screening and validation of thesemarkers. In certain embodiments, the present devices and methods areparticularly useful when coupled with data processing system capable ofpattern recognition for such purposes.

In certain embodiments, the devices and methods of the present inventionare also advantageous to create large sample dataset for refining thealgorithms for pattern recognition through machine learning and/or othermethodologies.

EBC-4. EBC Collection and Analysis without Spacers

Another aspect of the present invention is to provide devices andmethods for collecting and analyzing vapor condensate using theaforementioned collection plate and cover plate but without spacers.

In some embodiments of the present invention, the spacers that are usedto regulate the sample or a relevant volume of the EBC sample arereplaced by (a) positioning sensors that can measure the plate innerspacing, and/or (b) devices that can control the plate positions andmove the plates into a desired plate inner spacing based on theinformation provided the sensors. In some embodiment, all the spacersare replaced by translation stage, monitoring sensors and feedbacksystem.

In some embodiments, the collection plate and the cover plate compriseno spacers at all, and the EBC sample is compressed by the two platesinto a thin layer, the thickness of which is regulated by the spacingbetween the inner surfaces of the plates (the plate spacing).

A4. A device for collecting EBC without spacers, comprises:

a first plate and a second plate, wherein:

-   -   i. the plates are movable relative to each other into different        configurations, and one or both plates are flexible;    -   ii. both plates comprise a sample contact area on the respective        surface of each plate for contacting EBC sample;        -   wherein one of the configurations is an open configuration,            in which: the two plates are separated apart, and the EBC            sample is deposited on one or both of the plates from a            subject; and        -   wherein another of the configurations is a closed            configuration which is configured after the EBC sample            deposition in the open configuration; and in the closed            configuration: at least part of the EBC sample is compressed            by the two plates into a thin layer, wherein the thin layer            is in contact with and confined by the inner surfaces of the            two plates.

A5. A method of collecting EBC without spacers, comprises the steps:

(a) obtaining a collection plate and a cover plate of paragraph A4;

(b) depositing, when the plates are configured in the openconfiguration, an EBC sample by exhaling breath from a subject towardthe collection plate, wherein the exhaled breath condensates on acollection surface of the collection plate to form droplets and/orpuddles that have different lateral sizes and different heights,depending upon the surface wetting properties of the collection surface;

(c) after (b), bringing the cover plate over the collection surface andthen bringing the two plates into a closed configuration by pressing theplates, wherein at the closed configuration:

-   -   (i) at least a part of the EBC sample is between the cover plate        and the collection plate, and a relevant area of the collection        surface of the collection plate is covered by the cove plate;        and    -   (ii) in the relevant area, a substantial number or all of the        droplets or puddles formed in step (b) at the open configuration        merge into puddle(s) that (1) have much larger lateral size but        in a smaller number than the open configuration and (2) touch        both inner surfaces of the cover plate and the collection plate,        thereby the thickness of the puddle(s) is confined by the inner        surfaces of the plates and equal to the spacing between the        inner surfaces, and the total surface area of the deposited EBC        exposed to the ambient is significantly reduced; and

wherein the plate spacing is the spacing between the inner surfaces ofthe cover plate and the collection plate, the relevant area is a portionor entire surface of the collection surface, and the collection surfaceis a portion or entire surface of the collection plate.

A6. A method of analyzing EBC without spacers, comprises the steps:

(a) obtaining a collection plate and a cover plate of paragraph A4;

(b) depositing, when the plates are configured in the openconfiguration, an EBC sample by exhaling breath from a subject towardthe collection plate, wherein the exhaled breath condensates on acollection surface of the collection plate to form droplets and/orpuddles that have different lateral sizes and different heights,depending upon the surface wetting properties of the collection surface;

(c) after (b), bringing the cover plate over the collection surface andthen bringing the two plates into a closed configuration by pressing theplates, wherein at the closed configuration:

-   -   (i) at least a part of the EBC sample is between the cover plate        and the collection plate, and a relevant area of the collection        surface of the collection plate is covered by the cove plate;        and    -   (ii) in the relevant area, a substantial number or all of the        droplets or puddles formed in step (b) at the open configuration        merge into puddle(s) that (1) have much larger lateral size but        in a smaller number than the open configuration and (2) touch        both inner surfaces of the cover plate and the collection plate,        thereby the thickness of the puddle(s) is confined by the inner        surfaces of the plates and equal to the spacing between the        inner surfaces, and the total surface area of the deposited EBC        exposed to the ambient is significantly reduced; and

(d) analyzing the EBC,

wherein the plate spacing is the spacing between the inner surfaces ofthe cover plate and the collection plate, the relevant area is a portionor entire surface of the collection surface, and the collection surfaceis a portion or entire surface of the collection plate.

In some embodiments, it is unlikely to obtain a layer of highly uniformthickness without using the spacers as discussed in the foregoingsessions. However, it is still advantageous to use the device and methodof paragraphs A4-A5 for collecting and analyzing EBC sample, for it iseasy, rapid to handle, requires no professional training and a verysmall volume of sample.

In some embodiments, the analyzing step (d) of paragraph A6 comprisesdetermining the thickness of the collected EBC sample at the closedconfiguration after the formation of the thin layer during step (c). Insome embodiments, the thickness of the collected EBC sample at theclosed configuration is equal to the spacing between the inner surfacesof the two plates.

In some embodiments, measuring the spacing between the inner surfacescomprises the use of optical interference. The optical interference canuse multiple wavelength. For example, the light signal due to theinterference of a light reflected at the inner surface of the firstplate and the second plate oscillate with the wavelength of the light.From the oscillation, one can determine the spacing between the innersurfaces. To enhance the interference signal, one of the inner surfacesor both can be coated with light reflection material.

In some embodiments, measuring the spacing between the inner surfacescomprises taking optical imaging (e.g. taking a 2D (two-dimensional)/3D(three-dimensional) image of the sample and the image taking can bemultiple times with different viewing angles, different wavelength,different phase, and/or different polarization) and image processing.

In some embodiments, the analyzing step (d) of paragraph A6 comprisesmeasuring the volume of the collected EBC sample based on the lateralarea and the thickness of the thin layer that are determined after theformation of the thin layer during step (c).

In some embodiments, measuring the entire sample area or volumecomprises taking optical imaging (e.g. taking a 2D (two-dimensional)/3D(three-dimensional) image of the sample and the image taking can bemultiple times with different viewing angles, different wavelength,different phase, and/or different polarization) and image processing.The sample lateral area means the area in the direction approximatelyparallel to the first plate and the second plate. The 3D imaging can usethe method of fringe projection profilometry (FPP), which is one of themost prevalent methods for acquiring three-dimensional (3D) images ofobjects.

In some embodiments, the measuring of the sample area or volume byimaging comprises: (a) calibration of the image scale by using a sampleof the known area or volume (e.g., The imager is a smartphone and thedimensions of the image taken by the phone can be calibrated bycomparing an image of the a sample of known dimension taken the samephone); (b) comparison of the image with the scale markers (rulers)placed on or near the first plate and second plate (discussed furtherherein), and (c) a combination of thereof.

As used herein, light may include visible light, ultraviolet light,infrared light, and/or near infrared light. Light may includewavelengths in the range from 20 nm to 20,000 nm.

In some embodiments, the pressing during step (c) of paragraphs A5-A6 isperformed by human hand.

In some embodiments, the formation and properties of the thin layer isdependent on the pressing force applied during step (c) of paragraphsA5-A6 for bringing the two plates into the closed configuration (asdemonstrated in EBC-6). In some embodiments, the pressing force appliedduring step (c) of paragraphs A5-A6 is well adjusted for forming a thinlayer of EBC sample between the two plates that has prerequisiteparameters.

EBC-5. More Examples of EBC Collection and Analysis Experiments

Additional exemplary experimental testing and observation, andadditional preferred embodiments of the present invention are given.

All the exemplary experimental testing and demonstration of the presentinvention described in Section 4 (Examples) were performed under thefollowing conditions and share the following common observations.

Plates. Only one of the two plates of SiEBCA device, termed “X-Plate”,has the spacers fixed on the sample surface of the plate, and the otherplate, termed “the substrate plate”, has a planar surface and does nothave spacers. Unless particularly specified, the substrate plate wasused as the collection plate, and the X-plate was used as the coverplate. Various materials (including glass, PMMA (polymethacrylate), andPS (polystyrene)) for the plates and various plate thicknesses have beentested. The planar surface of the plates typically have surfaceroughness less than 30 nm.

Spacers. The spacers used on the X-Plate are rectangle pillars in aperiodic array with a fixed inner spacer distance (ISD) and uniformspacer height. The pillar spacers have a straight sidewall with a tiltangle from the normal less than 5 degree. Different spacer height, size,inter-spacer distance, shape, and materials are tested.

Fabrication of Spacers. The spacers are fabricated by nanoimprint on aplastic plate, where a mold is pressed directly into the plate. The moldwas fabricated by lithography and etching. Examples of the spacers onthe plate for SiEBCA. The spacers are fabricated by direct imprinting ofthe plastic plate surface using a mold, and has a dimension of width,length and height of 30 um, 40 um and 2 um.

EBC Sample deposition. All of the EBC samples were deposited on thecollection plates by having a human subject directly exhale toward thecollection plate which is placed within a few inches away from thesubject's mouth.

The EBC samples depositions were performed in standard room conditionswithout any special temperature control or dust filters. We found thatin our experiments, the dust does not affect to achieve thepredetermined final sample thickness over a large sample area, and atthe closed configuration the sample thickness over the non-dust area isregulated by the spacers. This demonstrated that the embodiments that weused for CROF performed to the results expected by the presentinvention.

Plate's surface wetting properties. Unless particularly specified, allthe sample surfaces (i.e. the inner surface that contacts a sample) ofthe plates are untreated. We have tested the wetting properties of theseuntreated surfaces as a function of the plate material (glass, PMMA, andPS), the surface structures (planar or with spacers, and the sample type(water, PBS buffer and blood), by dropping a small drop of sample on theplate surface and measuring the sample to the plate contact angle. Thewetting angles of the different surfaces for different samples werefound experimentally as follows: For the liquid of water, PBS, andblood, the contact angle is about 46 degree for untreated glass, 60degree for untreated PMMA surface, 60 degree for untreated PS(polystyrene) and about 61 degrees for untreated PMMA X-plate. Thereforethey are all hydrophilic. But the wetting property of these surfaces canchanged to either hydrophilic or hydrophobic by surface treatment. For agood vapor condensate collection, a hydrophilic surface is preferred,which will have, for a given amount of the condensation, smaller surfacearea by Hand-Press.

The present invention has performed various experiments in testing SiVCdevices and methods, which are partially described in the PCT [Julia:Inset the info] (ESX-002PCT), which is incorporated herein for itsentirety. Some of the experiments are summaried below (which used thedevice with spacers), but the present invention has preformed additionalexperiments and found that these observations are working for (and canbused) the SiVC device without spacers.

In all the experiment of the present invention, human hand(s) were themeans to press a SiEBCA device (plates) into a closed configuration,which is referred as “hand-pressing”.

Self-Holding. Self-holding means that after a SiEBCA device (plates) iscompressed into a closed configuration by an external force (e.g. theforce from hand) and after the external force is removed, the SiEBCAdevice can hold, on its own, the sample thickness unchanged. We observedthat in all the experiments in the Sec., all the SiEBCA devices andprocess (unless particularly specified) can self-hold, as demonstratedin the experiments. Our other experimental test showed that as long asone of the plate is hydrophilic, the SiEBCA plates can self-hold.

Drying speed with treated and untreated collection plates. For bothexperiments using treated and untreated collection plates, the dryingspeed of EBC was also calculated, which is defined as the retractionlength per unit time of the edge of the liquid sample in the X-device atthe closed configuration. The calculation shows that with the treatedPMMA collection plate, the liquid sample dried at a slower speed (74um/min) as compared to with the untreated PMMA collection plate (117um/min).

Surface treatment. The surface treatment of the PMMA plate was performedwith oxygen plasma, followed by deposition of 10 nm silicon oxide. Insome embodiments, the treatment was performed chemically usingtrimethoxysilane.

AA2. The device of embodiment AA0 or AA1, wherein the device furthercomprises a dry reagent coated on one or both of the plates.AA3. The device of any prior embodiment, wherein the device furthercomprises, on one or both plates, a dry binding site that has apredetermined area, wherein the dry binding site binds to andimmobilizes an analyte in the sample.AA4. The device of any prior embodiment, wherein the device furthercomprises, on one or both plates, a releasable dry reagent and a releasetime control material that delays the time that the releasable dryregent is released into the sample.AA5. The device of embodiment 4, wherein the release time controlmaterial delays the time that the dry regent starts is released into thesample by at least 3 seconds.AA6. The device of any prior embodiment, wherein the device furthercomprises, on one or both plates, one or a plurality of dry bindingsites and/or one or a plurality of reagent sites.AA7. The device of any prior embodiment, wherein the sample is exhalebreath condensate.AA8. The device of any prior embodiment, wherein the sample is a vaporfrom a biological sample, an environmental sample, a chemical sample, orclinical sample.AA9. The device of any prior embodiment, wherein the analyte comprises amolecule (e.g., a protein, peptides, DNA, RNA, nucleic acid, or othermolecules), cells, tissues, viruses, and nanoparticles with differentshapes.AA10. The device of any prior embodiment, wherein the analyte comprisesvolatile organic compounds (VOCs).AA11. The device of any prior embodiment, wherein the analyte comprisesnitrogen, oxygen, CO2, H2O, and inert gases.AA12. The device of any prior embodiment, wherein the analyte isstained.AA13. The device of any prior embodiment, wherein on one of the samplesurface, it further comprises an enclosure-spacer that encloses apartial or entire VC samples deposited on the collection plate.AA14. The device of any prior embodiment, wherein the highly uniformthickness has a value equal to or less than 0.5 um.AA15. The device of any prior embodiment, wherein the highly uniformthickness has a value in the range of 0.5 um to 1 um.AA16. The device of any prior embodiment, wherein the highly uniformthickness has a value in the range of 1 um to 2 um.AA17. The device of any prior embodiment, wherein the highly uniformthickness has a value in the range of 2 um to 10 um.AA18. The device of any prior embodiment, wherein the highly uniformthickness has a value in the range of 10 um to 20 um.AA19. The device of any prior embodiment, wherein the highly uniformthickness has a value in the range of 20 um to 30 um.AA20. The device of any prior embodiment, wherein the thickness of theat least a part of VC sample at the closed configuration is larger thanthe thickness of VC sample deposited on the collection plate at an openconfiguration.AA21. The device of any prior embodiment, wherein the thickness of theat least a part of VC sample at the closed configuration is less thanthe thickness of VC sample deposited on the collection plate at an openconfiguration.AA22. The device of any prior device embodiment, wherein the spacers arefixed on a plate by directly embossing the plate or injection molding ofthe plate.AA23. The device of any prior device embodiment, wherein the materialsof the plate and the spacers are selected from polystyrene, PMMA, PC,COC, COP, or another plastic.AA24. The device of any prior embodiment, wherein the inter-spacerspacing is in the range of 1 um to 200 um.AA25. The device of any prior embodiment, wherein the inter-spacerspacing is in the range of 200 um to 1000 um.AA26. The device of any prior embodiment, wherein the VC sample is anexhaled breath condensate from a human or an animal.AA27. The device of any prior embodiment, wherein the spacers regulatingthe layer of uniform thickness have a filling factor of at least 1%,wherein the filling factor is the ratio of the spacer area in contactwith the layer of uniform thickness to the total plate area in contactwith the layer of uniform thickness.AA28. The device of any prior embodiment, wherein for spacers regulatingthe layer of uniform thickness, the Young's modulus of the spacers timesthe filling factor of the spacers is equal to or larger than 10 MPa,wherein the filling factor is the ratio of the spacer area in contactwith the layer of uniform thickness to the total plate area in contactwith the layer of uniform thickness.AA29. The device of any prior embodiment, wherein for a flexible plate,the thickness of the flexible plate times the Young's modulus of theflexible plate is in the range 60 to 750 GPa-um.AA30. The device of any prior embodiment, wherein for a flexible plate,the fourth power of the inter-spacer-distance (ISD) divided by thethickness of the flexible plate (h) and the Young's modulus (E) of theflexible plate, ISD4/(hE), is equal to or less than 106 um3/GPa,AA31. The device of any prior paragraph, wherein one or both platescomprises a location marker, either on a surface of or inside the plate,that provides information of a location of the plate.AA32. The device of any prior paragraph, wherein one or both platescomprises a scale marker, either on a surface of or inside the plate,that provides information of a lateral dimension of a structure of thesample and/or the plate.AA33. The device of any prior embodiment, wherein one or both platescomprises an imaging marker, either on surface of or inside the plate,that assists an imaging of the sample.AA34. The device of any prior embodiment, wherein the spacers functionas a location marker, a scale marker, an imaging marker, or anycombination of thereof.AA35. The device of any prior embodiment, wherein the average thicknessof the layer of uniform thickness is about equal to a minimum dimensionof an analyte in the sample.AA36. The device of any prior embodiment, wherein the inter-spacerdistance is in the range of 1 um to 50 um.AA37. The device of any prior embodiment, wherein the inter-spacerdistance is in the range of 50 um to 120 um.AA38. The device of any prior embodiment, wherein the inter-spacerdistance is in the range of 120 um to 200 um.AA39. The device of any prior embodiment, wherein the inter-spacerdistance is substantially periodic.AA40. The device of any prior embodiment, wherein the inter-spacerdistance is aperiodic.AA41. The device of any prior embodiment, wherein the spacers arepillars with a cross-sectional shape selected from round, polygonal,circular, square, rectangular, oval, elliptical, or any combination ofthe same.AA42. The device of any prior embodiment, wherein the spacers have apillar shape and have a substantially flat top surface, wherein, foreach spacer, the ratio of the lateral dimension of the spacer to itsheight is at least 1.AA43. The device of any prior embodiment, wherein each spacer has aratio of the lateral dimension of the spacer to its height at least 1.AA44. The device of any prior embodiment, wherein the minimum lateraldimension of spacer is less than or substantially equal to the minimumdimension of an analyte in the sample.AA45. The device of any prior embodiment, wherein the minimum lateraldimension of spacer is in the range of 0.5 um to 100 um.AA46. The device of any prior embodiment, wherein the minimum lateraldimension of spacer is in the range of 0.5 um to 10 um.AA47. The device of any prior embodiment, wherein the spacers have adensity of at least 100/mm².AA48. The device of any prior embodiment, wherein the spacers have adensity of at least 1000/mm².AA49. The device of any prior embodiment, wherein at least one of theplates is transparent.AA50. The device of any prior embodiment, wherein at least one of theplates is made from a flexible polymer.AA51. The device of any prior embodiment, wherein, for a pressure thatcompresses the plates, the spacers are not compressible and/or,independently, only one of the plates is flexible.AA52. The device of any of any prior embodiment, wherein the flexibleplate has a thickness in the range of 10 um to 200 um.AA53. The device of any prior embodiment, wherein the variation is lessthan 30%.AA54. The device of any prior embodiment, wherein the variation is lessthan 10%.AA55. The device of any prior embodiment, wherein the variation is lessthan 5%.AA56. The device of any prior embodiment, wherein the first and secondplates are connected and are configured to be changed from the openconfiguration to the closed configuration by folding the plates.AA57. The device of any prior embodiment, wherein the first and secondplates are connected by a hinge and are configured to be changed fromthe open configuration to the closed configuration by folding the platesalong the hinge.AA58. The device of any prior embodiment, wherein the first and secondplates are connected by a hinge that is a separate material to theplates, and are configured to be changed from the open configuration tothe closed configuration by folding the plates along the hinge.AA59. The device of any prior embodiment, wherein the first and secondplates are made in a single piece of material and are configured to bechanged from the open configuration to the closed configuration byfolding the plates.AA60. The device of any prior embodiment, wherein the layer of uniformthickness sample is uniform over a lateral area that is at least 100um².AA61. The device of any prior embodiment, wherein the layer of uniformthickness sample is uniform over a lateral area that is at least 1 mm².AA62. The device of any prior embodiment, wherein the device isconfigured to analyze the sample in 60 seconds or less.AA63. The device of any prior embodiment, wherein at the closedconfiguration, the final sample thickness device is configured toanalyze the sample in 60 seconds or less.AA64. The device of any prior embodiment, wherein the device furthercomprises, on one or both of the plates, one or a plurality ofamplification sites that are each capable of amplifying a signal fromthe analyte or a label of the analyte when the analyte or label iswithin 500 nm from an amplification site.AA65. The device of any prior embodiment, wherein at the closedconfiguration, the final sample thickness device is configured toanalyze the sample in 10 seconds or less.AA66. The device of any prior embodiment, wherein the dry binding sitecomprises a capture agent.AA67. The device of any prior embodiment, wherein the dry binding sitecomprises an antibody or nucleic acid.AA68. The device of any prior embodiment, wherein the releasable dryreagent is a labeled reagent.AA69. The device of any prior embodiment, wherein the releasable dryreagent is a fluorescently-labeled reagent.AA70. The device of any prior embodiment, wherein the releasable dryreagent is a fluorescently-labeled antibody.AA71. The device of any prior embodiment, wherein the first platefurther comprises, on its surface, a first predetermined assay site anda second predetermined assay site, wherein the distance between theedges of the assay site is substantially larger than the thickness ofthe uniform thickness layer when the plates are in the closed position,wherein at least a part of the uniform thickness layer is over thepredetermined assay sites, and wherein the sample has one or a pluralityof analytes that are capable of diffusing in the sample.AA72. The device of any prior embodiment, wherein the first plate has,on its surface, at least three analyte assay sites, and the distancebetween the edges of any two neighboring assay sites is substantiallylarger than the thickness of the uniform thickness layer when the platesare in the closed position, wherein at least a part of the uniformthickness layer is over the assay sites, and wherein the sample has oneor a plurality of analytes that are capable of diffusing in the sample.AA73. The device of any prior embodiment, wherein the first plate has,on its surface, at least two neighboring analyte assay sites that arenot separated by a distance that is substantially larger than thethickness of the uniform thickness layer when the plates are in theclosed position, wherein at least a part of the uniform thickness layeris over the assay sites, and wherein the sample has one or a pluralityof analytes that are capable of diffusing in the sample.AA74. The device of any prior embodiment, wherein the releasable dryreagent is a cell stain.AA75. The device of any prior embodiment, wherein the device furthercomprises a detector that is an optical detector for detecting anoptical signal.AA76. The device of any prior embodiment, wherein the device furthercomprises a detector that is an electrical detector for detecting anelectric signal.AA77. The device of any prior embodiment, wherein the device comprisesdiscrete spacers that are not fixed to any of the plates, wherein at theclosed configuration, the discrete spacers are between the innersurfaces of the two plates, and the thickness of the sample is confinedby the inner surfaces of the two plates, and regulated by the discretespacers and the plates.AA78. The device of any prior embodiment, wherein the device furthercomprises a binding site that has a chemical sensor that is made from amaterial selected from the group consisting of: silicon nanowire (SiNW); single-walled carbon nanotubes (SWCNT); random networks of carbonnanotubes (RN-CNTs); molecularly capped metal nanoparticles (MCNPs);metal oxide nanoparticles (MONPs); and chemically sensitive field-effecttransistors (CHEM-FETs).BB1. A system for rapidly analyzing a vapor condensation sample using amobile phone comprising:(a) a device of any prior AA embodiment;(b) a mobile communication device comprising:

-   i. one or a plurality of cameras for the detecting and/or imaging    the vapor condensate sample; and-   ii. electronics, signal processors, hardware and software for    receiving and/or processing the detected signal and/or the image of    the vapor condensate sample and for remote communication.    BB2. The system of any prior BB embodiment, wherein the system    further comprise a light source from either the mobile communication    device or an external source.    BB3. The system of any prior BB embodiment, wherein one of the    plates has a binding site that binds an analyte, wherein at least    part of the uniform sample thickness layer is over the binding site,    and is substantially less than the average lateral linear dimension    of the binding site.    BB4. The system of any prior BB embodiment, further comprising:    (d) a housing configured to hold the sample and to be mounted to the    mobile communication device.    BB5. The system of any prior BB embodiment, wherein the housing    comprises optics for facilitating the imaging and/or signal    processing of the sample by the mobile communication device, and a    mount configured to hold the optics on the mobile communication    device.    BB6. The system of any prior BB embodiment, wherein an element of    the optics in the housing is movable relative to the housing.    BB7. The system of any prior BB embodiment, wherein the mobile    communication device is configured to communicate test results to a    medical professional, a medical facility or an insurance company.    BB8. The system of any prior BB embodiment, wherein the mobile    communication device is further configured to communicate    information on the test and the subject with the medical    professional, medical facility or insurance company.    BB9. The system of any prior BB embodiment, wherein the mobile    communication device is further configured to communicate    information of the test to a cloud network, and the cloud network    process the information to refine the test results.    BB10. The system of any prior BB embodiment, wherein the mobile    communication device is further configured to communicate    information of the test and the subject to a cloud network, the    cloud network process the information to refine the test results,    and the refined test results will send back the subject.    BB11. The system of any prior BB embodiment, wherein the mobile    communication device is configured to receive a prescription,    diagnosis or a recommendation from a medical professional.    BB12. The system of any prior BB embodiment, wherein the mobile    communication device is configured with hardware and software to:    (a) capture an image of the sample;    (b) analyze a test location and a control location in in image; and    (c) compare a value obtained from analysis of the test location to a    threshold value that characterizes the rapid diagnostic test.    BB13. The system of any prior BB embodiment, wherein at least one of    the plates comprises a storage site in which assay reagents are    stored.    BB14. The system of any prior BB embodiment, at least one of the    cameras reads a signal from the CROF device.    BB15. The system of any prior BB embodiment, wherein the mobile    communication device communicates with the remote location via a    wifi or cellular network.    BB16. The system of any prior BB embodiment, wherein the mobile    communication device is a mobile phone.    CC1. A method for rapidly analyzing an analyte in a sample using a    mobile phone, comprising:    (a) depositing a sample on the device of any prior BB embodiment;    (b) assaying an analyte in the sample deposited on the device to    generate a result; and    (c) communicating the result from the mobile communication device to    a location remote from the mobile communication device.    CC2. The method of any prior CC embodiment, wherein the analyte    comprises a molecule (e.g., a protein, peptides, DNA, RNA, nucleic    acid, or other molecule), cells, tissues, viruses, and nanoparticles    with different shapes.    CC3. The method of any prior CC embodiment, wherein the analyte    comprises white blood cell, red blood cell and platelets.    CC4. The method of any prior CC embodiment, wherein the method    comprises:    analyzing the results at the remote location to provide an analyzed    result; and communicating the analyzed result from the remote    location to the mobile communication device.    CC5. The method of any prior CC embodiment, wherein the analysis is    done by a medical professional at a remote location.    CC6. The method of any prior CC embodiment, wherein the mobile    communication device receives a prescription, diagnosis or a    recommendation from a medical professional at a remote location.    CC7. The method of any prior CC embodiment, wherein the thickness of    the at least a part of VC sample at the closed configuration is    larger than the thickness of VC sample deposited on the collection    plate at an open configuration.    CC8. The method of any prior CC embodiment, wherein the thickness of    the at least a part of VC sample at the closed configuration is less    than the thickness of VC sample deposited on the collection plate at    an open configuration.    CC9. The method of any prior CC paragraph, wherein the assaying step    comprises detecting an analyte in the sample.    CC10. The method of any prior CC paragraph, wherein the analyte is a    biomarker.    CC11. The method of any prior CC embodiment, wherein the analyte is    a protein, nucleic acid, cell, or metabolite.    CC12. The method of any prior CC embodiment, wherein the assay done    in step (b) is a binding assay or a biochemical assay.    DD1. A method for analyzing an analyte in a vapor condensate sample    comprising: obtaining a device of any prior device embodiment;    depositing the vapor condensate sample onto one or both pates of the    device;    placing the plates in a closed configuration and applying an    external force over at least part of the plates; and    analyzing the analytes in the layer of uniform thickness while the    plates are the closed configuration.    DD2. The method of any prior DD embodiment, wherein the method    comprises:

(a) obtaining a sample;

(b) obtaining a first and second plates that are movable relative toeach other into different configurations, wherein each plate has asample contact surface that is substantially planar, one or both platesare flexible, and one or both of the plates comprise spacers that arefixed with a respective sample contacting surface, and wherein thespacers have:

-   -   i. a predetermined substantially uniform height,    -   ii. a shape of pillar with substantially uniform cross-section        and a flat top surface;    -   iii. a ratio of the width to the height equal or larger than        one;    -   iv. a predetermined constant inter-spacer distance that is in        the range of 10 um to 200 um;    -   v. a filling factor of equal to 1% or larger;

(c) depositing the sample on one or both of the plates when the platesare configured in an open configuration, wherein the open configurationis a configuration in which the two plates are either partially orcompletely separated apart and the spacing between the plates is notregulated by the spacers;

(d), after (c), using the two plates to compress at least part of thesample into a layer of substantially uniform thickness that is confinedby the sample contact surfaces of the plates, wherein the uniformthickness of the layer is regulated by the spacers and the plates, andhas an average value equal to or less than 30 um with a variation ofless than 10%, wherein the compressing comprises:

bringing the two plates together; and

conformable pressing, either in parallel or sequentially, an area of atleast one of the plates to press the plates together to a closedconfiguration, wherein the conformable pressing generates asubstantially uniform pressure on the plates over the at least part ofthe sample, and the pressing spreads the at least part of the samplelaterally between the sample contact surfaces of the plates, and whereinthe closed configuration is a configuration in which the spacing betweenthe plates in the layer of uniform thickness region is regulated by thespacers; and

(e) analyzing the in the layer of uniform thickness while the plates arethe closed configuration;

wherein the filling factor is the ratio of the spacer contact area tothe total plate area; wherein a conformable pressing is a method thatmakes the pressure applied over an

area is substantially constant regardless the shape variation of theouter surfaces of the plates; and

wherein the parallel pressing applies the pressures on the intended areaat the same time, and a sequential pressing applies the pressure on apart of the intended area and gradually move to other area.

DD3. The method of any prior DD embodiment, wherein the methodcomprises:

-   -   removing the external force after the plates are in the closed        configuration; and

imaging the analytes in the layer of uniform thickness while the platesare the closed configuration; and

-   -   counting a number of analytes or the labels in an area of the        image.        DD4. The method of any prior DD embodiment, wherein the method        comprises removing the external force after the plates are in        the closed configuration; and measuring optical signal in the        layer of uniform thickness while the plates are the closed        configuration.        DD5. The method of any prior DD embodiment, wherein the        inter-spacer distance is in the range of 20 um to 200 um.        DD6. The method of any prior DD embodiment, wherein the        inter-spacer distance is in the range of 5 um to 20 um.        DD7. The method of any prior DD embodiment, wherein a product of        the filling factor and the Young's modulus of the spacer is 2        MPa or larger.        DD8. The method of any prior DD embodiment, the surface        variation is less than 50 nm.        DD9. The method of any prior DD embodiment, further comprising a        step of calculating the concentration of an analyte in the        relevant volume of sample, wherein the calculation is based on        the relevant sample volume defined by the predetermined area of        the storage site, the uniform sample thickness at the closed        configuration, and the amount of target entity detected.        DD10. The method of any prior DD embodiment, wherein the        analyzing step comprise counting the ananlyte in the sample.        DD11. The method of any prior DD embodiment, wherein the imaging        and counting is done by:

i. illuminating the cells in the layer of uniform thickness;

ii. taking one or more images of the cells using a CCD or CMOS sensor;

iii. identifying cells in the image using a computer; and

iv. counting a number of cells in an area of the image.

DD12. The method of any prior DD embodiment, wherein the external forceis provided by human hand.DD13. The method of any prior DD embodiment, wherein it future comprisesa dry reagent coated on one or both plates.DD14. The method of any prior DD embodiment, wherein the layer ofuniform thickness sample has a thickness uniformity of up to +/−5%.DD15. The method of any prior DD embodiment, wherein the spacers arepillars with a cross-sectional shape selected from round, polygonal,circular, square, rectangular, oval, elliptical, or any combination ofthe same.DD16. The method of any prior DD embodiment, wherein the spacing betweenthe spacers is approximately the minimum dimension of an analyte.EE1. The method of any prior CC or DD embodiment, wherein one or bothplate sample contact surfaces comprises one or a plurality ofamplification sites that are each capable of amplifying a signal fromthe analyte or a label of the analyte when the analyte or label iswithin 500 nm from an amplification site.EE2. The method of any prior CC or DD embodiment, wherein the sample isexhale breath condensate.EE3. The method of any prior CC or DD embodiment, wherein the sample isa vapor from a biological sample, an environmental sample, a chemicalsample, or clinical sample.EE4. The method of any prior CC or DD embodiment, wherein the analytecomprises a molecule (e.g., a protein, peptides, DNA, RNA, nucleic acid,or other molecules), cells, tissues, viruses, and nanoparticles withdifferent shapes.EE5. The method of any prior CC or DD embodiment, wherein the analytecomprises volatile organic compounds (VOCs).EE6. The method of any prior CC or DD embodiment, wherein the analytecomprises nitrogen, oxygen, CO2, H2O, and inert gases.EE7. The method of any prior CC or DD embodiment, wherein the analyte isstained.EE8. The method of any prior CC or DD embodiment, wherein on one of thesample surface, it further comprises an enclosure-spacer that encloses apartial or entire VC samples deposited on the collection plate.EE9. The method of any prior CC or DD embodiment, wherein the highlyuniform thickness has a value equal to or less than 0.5 um.EE10. The method of any prior CC or DD embodiment, wherein the highlyuniform thickness has a value in the range of 0.5 um to 1 um.EE11. The method of any prior CC or DD embodiment, wherein the highlyuniform thickness has a value in the range of 1 um to 2 um.EE12. The method of any prior CC or DD embodiment, wherein the highlyuniform thickness has a value in the range of 2 um to 10 um.EE13. The method of any prior CC or DD embodiment, wherein the highlyuniform thickness has a value in the range of 10 um to 20 um.EE14. The method of any prior CC or DD embodiment, wherein the highlyuniform thickness has a value in the range of 20 um to 30 um.

Other Embodiments

The present invention includes a variety of embodiments, which can becombined in multiple ways as long as the various components do notcontradict one another. The embodiments should be regarded as a singleinvention file: each filing has other filing as the references and isalso referenced in its entirety and for all purpose, rather than as adiscrete independent. These embodiments include not only the disclosuresin the current file, but also the documents that are herein referenced,incorporated, or to which priority is claimed.

(1) Definitions

The terms used in describing the devices, systems, and methods hereindisclosed are defined in the current application, or in PCT Application(designating U.S.) Nos. PCT/US2016/045437 and PCT/US0216/051775, whichwere respectively filed on Aug. 10, 2016 and Sep. 14, 2016, U.S.Provisional Application No. 62/456,065, which was filed on Feb. 7, 2017,U.S. Provisional Application No. 62/426,065, which was filed on Feb. 8,2017, U.S. Provisional Application No. 62/456,504, which was filed onFeb. 8, 2017, all of which applications are incorporated herein in theirentireties for all purposes.

The terms “CROF Card (or card)”, “COF Card”, “QMAX-Card”, “Q-Card”,“CROF device”, “COF device”, “QMAX-device”, “CROF plates”, “COF plates”,and “QMAX-plates” are interchangeable, except that in some embodiments,the COF card does not comprise spacers; and the terms refer to a devicethat comprises a first plate and a second plate that are movablerelative to each other into different configurations (including an openconfiguration and a closed configuration), and that comprises spacers(except some embodiments of the COF card) that regulate the spacingbetween the plates. The term “X-plate” refers to one of the two platesin a CROF card, wherein the spacers are fixed to this plate. Moredescriptions of the COF Card, CROF Card, and X-plate are given in theprovisional application serial nos. 62/456,065, filed on Feb. 7, 2017,which is incorporated herein in its entirety for all purposes.

(2) Q-Card, Spacer and Uniform Sample Thickness

The devices, systems, and methods herein disclosed can include or useQ-cards, spacers, and uniform sample thickness embodiments for sampledetection, analysis, and quantification. In some embodiments, the Q-cardcomprises spacers, which help to render at least part of the sample intoa layer of high uniformity. The structure, material, function, variationand dimension of the spacers, as well as the uniformity of the spacersand the sample layer, are herein disclosed, or listed, described, andsummarized in PCT Application (designating U.S.) Nos. PCT/US2016/045437and PCT/US0216/051775, which were respectively filed on Aug. 10, 2016and Sep. 14, 2016, U.S. Provisional Application No. 62/456,065, whichwas filed on Feb. 7, 2017, U.S. Provisional Application No. 62/426,065,which was filed on Feb. 8, 2017, U.S. Provisional Application No.62/456,504, which was filed on Feb. 8, 2017, all of which applicationsare incorporated herein in their entireties for all purposes.

(3) Hinges, Opening Notches, Recessed Edge and Sliders

The devices, systems, and methods herein disclosed can include or useQ-cards for sample detection, analysis, and quantification. In someembodiments, the Q-card comprises hinges, notches, recesses, andsliders, which help to facilitate the manipulation of the Q card and themeasurement of the samples. The structure, material, function, variationand dimension of the hinges, notches, recesses, and sliders are hereindisclosed, or listed, described, and summarized in PCT Application(designating U.S.) Nos. PCT/US2016/045437 and PCT/US0216/051775, whichwere respectively filed on Aug. 10, 2016 and Sep. 14, 2016, U.S.Provisional Application No. 62/456,065, which was filed on Feb. 7, 2017,U.S. Provisional Application No. 62/426,065, which was filed on Feb. 8,2017, U.S. Provisional Application No. 62/456,504, which was filed onFeb. 8, 2017, all of which applications are incorporated herein in theirentireties for all purposes.

(4) Q-Card, Sliders, and Smartphone Detection System

The devices, systems, and methods herein disclosed can include or useQ-cards for sample detection, analysis, and quantification. In someembodiments, the Q-cards are used together with sliders that allow thecard to be read by a smartphone detection system. The structure,material, function, variation, dimension and connection of the Q-card,the sliders, and the smartphone detection system are herein disclosed,or listed, described, and summarized in PCT Application (designatingU.S.) Nos. PCT/US2016/045437 and PCT/US0216/051775, which wererespectively filed on Aug. 10, 2016 and Sep. 14, 2016, U.S. ProvisionalApplication No. 62/456,065, which was filed on Feb. 7, 2017, U.S.Provisional Application No. 62/426,065, which was filed on Feb. 8, 2017,U.S. Provisional Application No. 62/456,504, which was filed on Feb. 8,2017, all of which applications are incorporated herein in theirentireties for all purposes.

In some embodiments of QMAX, the sample contact area of one or both ofthe plates comprises a compressed open flow monitoring surfacestructures (MSS) that are configured to monitoring how much flow hasoccurred after COF. For examples, the MSS comprises, in someembodiments, shallow square array, which will cause friction to thecomponents (e.g. blood cells in a blood) in a sample. By checking thedistributions of some components of a sample, one can obtain informationrelated to a flow, under a COF, of the sample and its components.

The depth of the MSS can be 1/1000, 1/100, 1/100, ⅕, ½ of the spacerheight or in a range of any two values, and in either protrusion or wellform.

(5) Detection Methods

The devices, systems, and methods herein disclosed can include or beused in various types of detection methods. The detection methods areherein disclosed, or listed, described, and summarized in PCTApplication (designating U.S.) Nos. PCT/US2016/045437 andPCT/US0216/051775, which were respectively filed on Aug. 10, 2016 andSep. 14, 2016, U.S. Provisional Application No. 62/456,065, which wasfiled on Feb. 7, 2017, U.S. Provisional Application No. 62/426,065,which was filed on Feb. 8, 2017, U.S. Provisional Application No.62/456,504, which was filed on Feb. 8, 2017, all of which applicationsare incorporated herein in their entireties for all purposes.

(6) Labels

The devices, systems, and methods herein disclosed can employ varioustypes of labels that are used for analytes detection. The labels areherein disclosed, or listed, described, and summarized in PCTApplication (designating U.S.) Nos. PCT/US2016/045437 andPCT/US0216/051775, which were respectively filed on Aug. 10, 2016 andSep. 14, 2016, U.S. Provisional Application No. 62/456,065, which wasfiled on Feb. 7, 2017, U.S. Provisional Application No. 62/426,065,which was filed on Feb. 8, 2017, U.S. Provisional Application No.62/456,504, which was filed on Feb. 8, 2017, all of which applicationsare incorporated herein in their entireties for all purposes.

(7) Analytes

The devices, systems, and methods herein disclosed can be applied tomanipulation and detection of various types of analytes (includingbiomarkers). The analytes and are herein disclosed, or listed,described, and summarized in PCT Application (designating U.S.) Nos.PCT/US2016/045437 and PCT/US0216/051775, which were respectively filedon Aug. 10, 2016 and Sep. 14, 2016, U.S. Provisional Application No.62/456,065, which was filed on Feb. 7, 2017, U.S. ProvisionalApplication No. 62/426,065, which was filed on Feb. 8, 2017, U.S.Provisional Application No. 62/456,504, which was filed on Feb. 8, 2017,all of which applications are incorporated herein in their entiretiesfor all purposes.

(8) Applications (Field and Samples)

The devices, systems, and methods herein disclosed can be used forvarious applications (fields and samples). The applications are hereindisclosed, or listed, described, and summarized in PCT Application(designating U.S.) Nos. PCT/US2016/045437 and PCT/US0216/051775, whichwere respectively filed on Aug. 10, 2016 and Sep. 14, 2016, U.S.Provisional Application No. 62/456,065, which was filed on Feb. 7, 2017,U.S. Provisional Application No. 62/426,065, which was filed on Feb. 8,2017, U.S. Provisional Application No. 62/456,504, which was filed onFeb. 8, 2017, all of which applications are incorporated herein in theirentireties for all purposes.

(9) Cloud

The devices, systems, and methods herein disclosed can employ cloudtechnology for data transfer, storage, and/or analysis. The relatedcloud technologies are herein disclosed, or listed, described, andsummarized in PCT Application (designating U.S.) Nos. PCT/US2016/045437and PCT/US0216/051775, which were respectively filed on Aug. 10, 2016and Sep. 14, 2016, U.S. Provisional Application No. 62/456,065, whichwas filed on Feb. 7, 2017, U.S. Provisional Application No. 62/426,065,which was filed on Feb. 8, 2017, U.S. Provisional Application No.62/456,504, which was filed on Feb. 8, 2017, all of which applicationsare incorporated herein in their entireties for all purposes.

Aspects:

-   -   1. A device for collecting and analyzing vapor condensate (VC)        sample, comprising:        -   a. a collection plate and a cover plate, wherein:        -   the plates are movable relative to each other into different            configurations;            -   i. one or both plates are flexible; and            -   ii. each of the plates has, on its inner respective                surface, a sample contact area for contacting a vapor                condensate (VC) sample that contains an analyte;            -   iii. wherein one of the configurations is an open                configuration, in which:                -   the two plates are either completely or partially                    separated apart, and the VC sample is deposited on                    one or both of the plates; and                -   wherein another of the configurations is a closed                    configuration which is configured after the VC                    sample deposition in the open configuration;                -   and in the closed configuration: at least a part of                    the VC sample is between the two plates and in                    contact with the two plates, and has a thickness                    that (a) is regulated by the two sample contact                    surfaces of the plates without using spacers,                    and (b) is equal to or less than 30 um with a small                    variation.    -   2. A device for collecting and analyzing vapor condensate (VC)        sample, comprising:        -   a. a collection plate and a cover plate, wherein:        -   the plates are movable relative to each other into different            configurations;        -   one or both plates are flexible; and        -   each of the plates has, on its respective surface, a sample            contact area for contacting a vapor condensate (VC) sample            that contains an analyte;            -   i. wherein one of the configurations is an open                configuration, in which: the two plates are either                completely or partially separated apart, and the VC                sample is deposited on one or both of the plates; and            -   ii. wherein another of the configurations is a closed                configuration which is configured after the VC sample                deposition in the open configuration; and in the closed                configuration: at least a part of the VC sample is                between the two plates and in contact with the two                plates, and has a thickness that is regulated by the                plate spacing without using spacers.    -   3. A device for collecting and analyzing vapor condensate (VC)        sample, comprising:        -   a. a collection plate, a cover plate, and spacers, wherein:            -   i. the plates are movable relative to each other into                different configurations;            -   ii. one or both plates are flexible;            -   iii. each of the plates has, on its respective inner                surface, a sample contact area for contacting a vapor                condensate (VC) sample that contains an analyte;            -   iv. the spacers are fixed to the respective inner                surface of one or both of the plates, and have a                predetermined substantially uniform height and a                predetermined constant inter-spacer distance and wherein                at least one of the spacers is inside the sample contact                area;            -   v. wherein one of the configurations is an open                configuration, in which: the two plates are either                completely or partially separated apart, the spacing                between the plates is not regulated by the spacers, and                the VC sample is deposited on one or both of the plates;                and            -   vi. wherein another of the configurations is a closed                configuration which is configured after the VC sample                deposition in the open configuration; and in the closed                configuration: at least a part of the VC sample is                between the two plates and in contact with the two                plates, and has a highly uniform thickness that is                regulated by the spacers and the two sample contact                surfaces of the plates and is equal to or less than 30                um.    -   4. A system is provided herein for rapidly analyzing a vapor        condensate sample using a mobile phone comprising:        -   a. a device of any prior Aspects;        -   b. a mobile communication device comprising:            -   i. one or a plurality of cameras for the detecting                and/or imaging the vapor condensate sample; and            -   ii. electronics, signal processors, hardware and                software for receiving and/or processing the detected                signal and/or the image of the vapor condensate sample                and for remote communication.    -   5. The system of any prior Aspect, further comprising a light        source from either the mobile communication device or an        external source.    -   6. A method is provided herein for analyzing an analyte in a        vapor condensate sample comprising:        -   a. obtaining a device of any prior device Aspect;        -   b. depositing the vapor condensate sample onto one or both            pates of the device;        -   c. placing the plates in a closed configuration and applying            an external force over at least part of the plates; and        -   d. analyzing the analyts in the layer of uniform thickness            while the plates are the closed configuration.    -   7. The device of any prior Aspect, wherein the device is        configured to analyze the sample in 60 seconds or less.    -   8. The device of any prior Aspect, wherein, at the closed        configuration, the final sample thickness is configured to        analyze the sample in 60 seconds or less.    -   9. The device of any prior Aspect, wherein the device further        comprises a dry reagent coated on one or both of the plates.    -   10. The device of any prior Aspect, wherein the device further        comprises, on one or both plates, a dry binding site that has a        predetermined area, wherein the dry binding site binds to and        immobilizes an analyte in the sample.    -   11. The device of any prior Aspect, wherein the device further        comprises, on one or both plates, a releasable dry reagent and a        release time control material that delays the time that the        releasable dry regent is released into the sample.    -   12. The device of Aspect 11, wherein the release time control        material delays the time that the dry regent starts is released        into the sample by at least 3 seconds.    -   13. The device of any prior Aspect, wherein the device further        comprises, on one or both plates, one or a plurality of dry        binding sites and/or one or a plurality of reagent sites.    -   14. The device of any prior Aspect, wherein the sample is exhale        breath condensate.    -   15. The device of any prior Aspect, wherein the sample is a        vapor from a biological sample, an environmental sample, a        chemical sample, or clinical sample.    -   16. The device of any prior Aspect, wherein the analyte        comprises a molecule (e.g., a protein, peptides, DNA, RNA,        nucleic acid, or other molecules), cells, tissues, viruses, and        nanoparticles with different shapes.    -   17. The device of any prior Aspect, wherein the analyte        comprises volatile organic compounds (VOCs).    -   18. The device of any prior Aspect, wherein the analyte        comprises nitrogen, oxygen, CO2, H2O, and inert gases.    -   19. The device of any prior Aspect, wherein the analyte is        stained.    -   20. The device of any prior Aspects, wherein on one of the        sample surface, it further comprises an enclosure-spacer that        encloses a partial or entire VC samples deposited on the        collection plate.    -   21. The device of any prior Aspect, wherein the highly uniform        thickness has a value equal to or less than 0.5 um.    -   22. The device of any prior Aspect, wherein the highly uniform        thickness has a value in the range of 0.5 um to 1 um.    -   23. The device of any prior Aspect, wherein the highly uniform        thickness has a value in the range of 1 um to 2 um.    -   24. The device of any prior Aspect, wherein the highly uniform        thickness has a value in the range of 2 um to 10 um.    -   25. The device of any prior Aspect, wherein the highly uniform        thickness has a value in the range of 10 um to 20 um.    -   26. The device of any prior Aspect, wherein the highly uniform        thickness has a value in the range of 20 um to 30 um.    -   27. The device of any prior Aspect, wherein the thickness of the        at least a part of VC sample at the closed configuration is        larger than the thickness of VC sample deposited on the        collection plate at an open configuration.    -   28. The device of any prior Aspect, wherein the thickness of the        at least a part of VC sample at the closed configuration is less        than the thickness of VC sample deposited on the collection        plate at an open configuration.    -   29. The device of any prior device Aspect, wherein the spacers        are fixed on a plate by directly embossing the plate or        injection molding of the plate.    -   30. The device of any prior device Aspect, wherein the materials        of the plate and the spacers are selected from polystyrene,        PMMA, PC, COC, COP, or another plastic.    -   31. The device of any prior Aspect, wherein the inter-spacer        spacing is in the range of 1 um to 200 um.    -   32. The device of any prior Aspect, wherein the inter-spacer        spacing is in the range of 200 um to 1000 um.    -   33. The device of any prior Aspect, wherein the VC sample is an        exhaled breath condensate from a human or an animal.    -   34. The device of any prior Aspect, wherein the spacers        regulating the layer of uniform thickness have a filling factor        of at least 1%, wherein the filling factor is the ratio of the        spacer area in contact with the layer of uniform thickness to        the total plate area in contact with the layer of uniform        thickness.    -   35. The device of any prior Aspect, wherein for spacers        regulating the layer of uniform thickness, the Young's modulus        of the spacers times the filling factor of the spacers is equal        to or larger than 10 MPa, wherein the filling factor is the        ratio of the spacer area in contact with the layer of uniform        thickness to the total plate area in contact with the layer of        uniform thickness.    -   36. The device of any prior Aspect, wherein for a flexible        plate, the thickness of the flexible plate times the Young's        modulus of the flexible plate is in the range 60 to 750 GPa-um.    -   37. The device of any prior Aspect, wherein for a flexible        plate, the fourth power of the inter-spacer-distance (ISD)        divided by the thickness of the flexible plate (h) and the        Young's modulus (E) of the flexible plate, ISD4/(hE), is equal        to or less than 106 um3/GPa,    -   38. The device of any prior Aspect, wherein one or both plates        comprises a location marker, either on a surface of or inside        the plate, that provides information of a location of the plate.    -   39. The device of any prior Aspect, wherein one or both plates        comprises a scale marker, either on a surface of or inside the        plate, that provides information of a lateral dimension of a        structure of the sample and/or the plate.    -   40. The device of any prior Aspect, wherein one or both plates        comprises an imaging marker, either on surface of or inside the        plate, that assists an imaging of the sample.    -   41. The device of any prior Aspect, wherein the spacers function        as a location marker, a scale marker, an imaging marker, or any        combination of thereof.    -   42. The device of any prior Aspect, wherein the average        thickness of the layer of uniform thickness is about equal to a        minimum dimension of an analyte in the sample.    -   43. The device of any prior Aspect, wherein the inter-spacer        distance is in the range of 1 um to 50 um.    -   44. The device of any prior Aspect, wherein the inter-spacer        distance is in the range of 50 um to 120 um.    -   45. The device of any prior Aspect, wherein the inter-spacer        distance is in the range of 120 um to 200 um.    -   46. The device of any prior Aspect, wherein the inter-spacer        distance is substantially periodic.    -   47. The device of any prior Aspect, wherein the inter-spacer        distance is aperiodic.    -   48. The device of any prior Aspect, wherein the spacers are        pillars with a cross-sectional shape selected from round,        polygonal, circular, square, rectangular, oval, elliptical, or        any combination of the same.    -   49. The device of any prior Aspect, wherein the spacers have a        pillar shape and have a substantially flat top surface, wherein,        for each spacer, the ratio of the lateral dimension of the        spacer to its height is at least 1.    -   50. The device of any prior Aspect, wherein each spacer has a        ratio of the lateral dimension of the spacer to its height at        least 1.    -   51. The device of any prior Aspect, wherein the minimum lateral        dimension of spacer is less than or substantially equal to the        minimum dimension of an analyte in the sample.    -   52. The device of any prior Aspect, wherein the minimum lateral        dimension of spacer is in the range of 0.5 um to 100 um.    -   53. The device of any prior Aspect, wherein the minimum lateral        dimension of spacer is in the range of 0.5 um to 10 um.    -   54. The device of any prior Aspect, wherein the spacers have a        density of at least 100/mm².    -   55. The device of any prior Aspect, wherein the spacers have a        density of at least 1000/mm².    -   56. The device of any prior Aspect, wherein at least one of the        plates is transparent.    -   57. The device of any prior Aspect, wherein at least one of the        plates is made from a flexible polymer.    -   58. The device of any prior Aspect, wherein, for a pressure that        compresses the plates, the spacers are not compressible and/or,        independently, only one of the plates is flexible.    -   59. The device of any of any prior Aspect, wherein the flexible        plate has a thickness in the range of 10 um to 200 um.    -   60. The device of any prior Aspect, wherein the variation is        less than 30%.    -   61. The device of any prior Aspect, wherein the variation is        less than 10%.    -   62. The device of any prior Aspect, wherein the variation is        less than 5%.    -   63. The device of any prior Aspect, wherein the first and second        plates are connected and are configured to be changed from the        open configuration to the closed configuration by folding the        plates.    -   64. The device of any prior Aspect, wherein the first and second        plates are connected by a hinge and are configured to be changed        from the open configuration to the closed configuration by        folding the plates along the hinge.    -   65. The device of any prior Aspect, wherein the first and second        plates are connected by a hinge that is a separate material to        the plates, and are configured to be changed from the open        configuration to the closed configuration by folding the plates        along the hinge.    -   66. The device of any prior Aspect, wherein the first and second        plates are made in a single piece of material and are configured        to be changed from the open configuration to the closed        configuration by folding the plates.    -   67. The device of any prior Aspect, wherein the layer of uniform        thickness sample is uniform over a lateral area that is at least        100 um′.    -   68. The device of any prior Aspect, wherein the layer of uniform        thickness sample is uniform over a lateral area that is at least        1 mm².    -   69. The device of any prior Aspect, wherein the device is        configured to analyze the sample in 60 seconds or less.    -   70. The device of any prior Aspect, wherein at the closed        configuration, the final sample thickness device is configured        to analyze the sample in 60 seconds or less.    -   71. The device of any prior Aspect, wherein the device further        comprises, on one or both of the plates, one or a plurality of        amplification sites that are each capable of amplifying a signal        from the analyte or a label of the analyte when the analyte or        label is within 500 nm from an amplification site.    -   72. The device of any prior Aspect, wherein at the closed        configuration, the final sample thickness device is configured        to analyze the sample in 10 seconds or less.    -   73. The device of any prior Aspect, wherein the dry binding site        comprises a capture agent.    -   74. The device of any prior Aspect, wherein the dry binding site        comprises an antibody or nucleic acid.    -   75. The device of any prior Aspect, wherein the releasable dry        reagent is a labeled reagent.    -   76. The device of any prior Aspect, wherein the releasable dry        reagent is a fluorescently-labeled reagent.    -   77. The device of any prior Aspect, wherein the releasable dry        reagent is a fluorescently-labeled antibody.    -   78. The device of any prior Aspect, wherein the first plate        further comprises, on its surface, a first predetermined assay        site and a second predetermined assay site, wherein the distance        between the edges of the assay site is substantially larger than        the thickness of the uniform thickness layer when the plates are        in the closed position, wherein at least a part of the uniform        thickness layer is over the predetermined assay sites, and        wherein the sample has one or a plurality of analytes that are        capable of diffusing in the sample.    -   79. The device of any prior Aspect, wherein the first plate has,        on its surface, at least three analyte assay sites, and the        distance between the edges of any two neighboring assay sites is        substantially larger than the thickness of the uniform thickness        layer when the plates are in the closed position, wherein at        least a part of the uniform thickness layer is over the assay        sites, and wherein the sample has one or a plurality of analytes        that are capable of diffusing in the sample.    -   80. The device of any prior Aspect, wherein the first plate has,        on its surface, at least two neighboring analyte assay sites        that are not separated by a distance that is substantially        larger than the thickness of the uniform thickness layer when        the plates are in the closed position, wherein at least a part        of the uniform thickness layer is over the assay sites, and        wherein the sample has one or a plurality of analytes that are        capable of diffusing in the sample.    -   81. The device of any prior Aspect, wherein the releasable dry        reagent is a cell stain.    -   82. The device of any prior Aspect, wherein the device further        comprises a detector that is an optical detector for detecting        an optical signal.    -   83. The device of any prior Aspect, wherein the device further        comprises a detector that is an electrical detector for        detecting an electric signal.    -   84. The device of any prior Aspect, wherein the device comprises        discrete spacers that are not fixed to any of the plates,        wherein at the closed configuration, the discrete spacers are        between the inner surfaces of the two plates, and the thickness        of the sample is confined by the inner surfaces of the two        plates, and regulated by the discrete spacers and the plates.    -   85. The device of any prior Aspect, wherein the device further        comprises a binding site that has a chemical sensor that is made        from a material selected from the group consisting of: silicon        nanowire (Si NW); single-walled carbon nanotubes (SWCNT); random        networks of carbon nanotubes (RN-CNTs); molecularly capped metal        nanoparticles (MCNPs); metal oxide nanoparticles (MONPs); and        chemically sensitive field-effect transistors (CHEM-FETs).    -   86. A system for rapidly analyzing a vapor condensation sample        using a mobile phone comprising:        -   a. a device of any prior AA embodiment;        -   b. a mobile communication device comprising:            -   i. one or a plurality of cameras for the detecting                and/or imaging the vapor condensate sample; and            -   ii. electronics, signal processors, hardware and                software for receiving and/or processing the detected                signal and/or the image of the vapor condensate sample                and for remote communication.    -   87. The system of any prior Aspect, wherein the system further        comprise a light source from either the mobile communication        device or an external source.    -   88. The system of any prior Aspect, wherein one of the plates        has a binding site that binds an analyte, wherein at least part        of the uniform sample thickness layer is over the binding site,        and is substantially less than the average lateral linear        dimension of the binding site.    -   89. The system of any prior Aspect, further comprising:    -   (d) a housing configured to hold the sample and to be mounted to        the mobile communication device.    -   90. The system of any prior Aspect, wherein the housing        comprises optics for facilitating the imaging and/or signal        processing of the sample by the mobile communication device, and        a mount configured to hold the optics on the mobile        communication device.    -   91. The system of any prior Aspect, wherein an element of the        optics in the housing is movable relative to the housing.    -   92. The system of any prior Aspect, wherein the mobile        communication device is configured to communicate test results        to a medical professional, a medical facility or an insurance        company.    -   93. The system of any prior Aspect, wherein the mobile        communication device is further configured to communicate        information on the test and the subject with the medical        professional, medical facility or insurance company.    -   94. The system of any prior Aspect, wherein the mobile        communication device is further configured to communicate        information of the test to a cloud network, and the cloud        network process the information to refine the test results.    -   95. The system of any prior Aspect, wherein the mobile        communication device is further configured to communicate        information of the test and the subject to a cloud network, the        cloud network process the information to refine the test        results, and the refined test results will send back the        subject.    -   96. The system of any prior Aspect, wherein the mobile        communication device is configured to receive a prescription,        diagnosis or a recommendation from a medical professional.    -   97. The system of any prior Aspect, wherein the mobile        communication device is configured with hardware and software        to:        -   a. capture an image of the sample;        -   b. analyze a test location and a control location in in            image; and        -   c. compare a value obtained from analysis of the test            location to a threshold value that characterizes the rapid            diagnostic test.    -   98. The system of any prior Aspect, wherein at least one of the        plates comprises a storage site in which assay reagents are        stored.    -   99. The system of any prior Aspect, at least one of the cameras        reads a signal from the CROF device.    -   100. The system of any prior Aspect, wherein the mobile        communication device communicates with the remote location via a        wifi or cellular network.    -   101. The system of any prior Aspect, wherein the mobile        communication device is a mobile phone.    -   102. A method for rapidly analyzing an analyte in a sample using        a mobile phone, comprising:        -   a. depositing a sample on the device of any prior BB            embodiment;        -   b. assaying an analyte in the sample deposited on the device            to generate a result; and        -   c. communicating the result from the mobile communication            device to a location remote from the mobile communication            device.    -   103. The method of any prior Aspect, wherein the analyte        comprises a molecule (e.g., a protein, peptides, DNA, RNA,        nucleic acid, or other molecule), cells, tissues, viruses, and        nanoparticles with different shapes.    -   104. The method of any prior Aspect, wherein the analyte        comprises white blood cell, red blood cell and platelets.    -   105. The method of any prior Aspect, wherein the method        comprises:        -   a. analyzing the results at the remote location to provide            an analyzed result; and communicating the analyzed result            from the remote location to the mobile communication device.    -   106. The method of any prior Aspect, wherein the analysis is        done by a medical professional at a remote location.    -   107. The method of any prior Aspect, wherein the mobile        communication device receives a prescription, diagnosis or a        recommendation from a medical professional at a remote location.    -   108. The method of any prior Aspect, wherein the thickness of        the at least a part of VC sample at the closed configuration is        larger than the thickness of VC sample deposited on the        collection plate at an open configuration.    -   109. The method of any prior Aspect, wherein the thickness of        the at least a part of VC sample at the closed configuration is        less than the thickness of VC sample deposited on the collection        plate at an open configuration.    -   110. The method of any prior Aspect, wherein the assaying step        comprises detecting an analyte in the sample.    -   111. The method of any prior Aspect, wherein the analyte is a        biomarker.    -   112. The method of any prior Aspect, wherein the analyte is a        protein, nucleic acid, cell, or metabolite.    -   113. The method of any prior Aspect, wherein the assay done in        step (b) is a binding assay or a biochemical assay.    -   114. A method for analyzing an analyte in a vapor condensate        sample comprising:        -   a. obtaining a device of any prior device Aspect;        -   b. depositing the vapor condensate sample onto one or both            pates of the device;        -   c. placing the plates in a closed configuration and applying            an external force over at least part of the plates; and        -   d. analyzing the analytes in the layer of uniform thickness            while the plates are the closed configuration.    -   115. The method of any prior Aspect, wherein the method        comprises:        -   a. obtaining a sample;        -   b. obtaining a first and second plates that are movable            relative to each other into different configurations,            wherein each plate has a sample contact surface that is            substantially planar, one or both plates are flexible, and            one or both of the plates comprise spacers that are fixed            with a respective sample contacting surface, and wherein the            spacers have:            -   i. a predetermined substantially uniform height,            -   ii. a shape of pillar with substantially uniform                cross-section and a flat top surface;            -   iii. a ratio of the width to the height equal or larger                than one;            -   iv. a predetermined constant inter-spacer distance that                is in the range of 10 um to 200 um;            -   v. a filling factor of equal to 1% or larger;        -   c. depositing the sample on one or both of the plates when            the plates are configured in an open configuration, wherein            the open configuration is a configuration in which the two            plates are either partially or completely separated apart            and the spacing between the plates is not regulated by the            spacers;        -   d. (d), after (c), using the two plates to compress at least            part of the sample into a layer of substantially uniform            thickness that is confined by the sample contact surfaces of            the plates, wherein the uniform thickness of the layer is            regulated by the spacers and the plates, and has an average            value equal to or less than 30 um with a variation of less            than 10%, wherein the compressing comprises:            -   i. bringing the two plates together; and            -   ii. conformable pressing, either in parallel or                sequentially, an area of at least one of the plates to                press the plates together to a closed configuration,                wherein the conformable pressing generates a                substantially uniform pressure on the plates over the at                least part of the sample, and the pressing spreads the                at least part of the sample laterally between the sample                contact surfaces of the plates, and wherein the closed                configuration is a configuration in which the spacing                between the plates in the layer of uniform thickness                region is regulated by the spacers; and        -   e. analyzing the in the layer of uniform thickness while the            plates are the closed configuration;            -   i. wherein the filling factor is the ratio of the spacer                contact area to the total plate area; wherein a                conformable pressing is a method that makes the pressure                applied over an area is substantially constant                regardless the shape variation of the outer surfaces of                the plates; and            -   ii. wherein the parallel pressing applies the pressures                on the intended area at the same time, and a sequential                pressing applies the pressure on a part of the intended                area and gradually move to other area.    -   116. The method of any prior Aspect, wherein the method        comprises:        -   a. removing the external force after the plates are in the            closed configuration; and imaging the analytes in the layer            of uniform thickness while the plates are the closed            configuration; and        -   b. counting a number of analytes or the labels in an area of            the image.    -   117. The method of any prior Aspect, wherein the method        comprises removing the external force after the plates are in        the closed configuration; and measuring optical signal in the        layer of uniform thickness while the plates are the closed        configuration.    -   118. The method of any prior Aspect, wherein the inter-spacer        distance is in the range of 20 um to 200 um.    -   119. The method of any prior Aspect, wherein the inter-spacer        distance is in the range of 5 um to 20 um.    -   120. The method of any prior Aspect, wherein a product of the        filling factor and the Young's modulus of the spacer is 2 MPa or        larger.    -   121. The method of any prior Aspect, the surface variation is        less than 50 nm.    -   122. The method of any prior Aspect, further comprising a step        of calculating the concentration of an analyte in the relevant        volume of sample, wherein the calculation is based on the        relevant sample volume defined by the predetermined area of the        storage site, the uniform sample thickness at the closed        configuration, and the amount of target entity detected.

123. The method of any prior Aspect, wherein the analyzing step comprisecounting the ananlyte in the sample.

-   -   124. The method of any prior Aspect, wherein the imaging and        counting is done by:        -   a. illuminating the cells in the layer of uniform thickness;        -   b. taking one or more images of the cells using a CCD or            CMOS sensor;        -   c. identifying cells in the image using a computer; and        -   d. counting a number of cells in an area of the image.    -   125. The method of any prior Aspect, wherein the external force        is provided by human hand.    -   126. The method of any prior Aspect, wherein it future comprises        a dry reagent coated on one or both plates.    -   127. The method of any prior Aspect, wherein the layer of        uniform thickness sample has a thickness uniformity of up to        +/−5%.    -   128. The method of any prior Aspect, wherein the spacers are        pillars with a cross-sectional shape selected from round,        polygonal, circular, square, rectangular, oval, elliptical, or        any combination of the same.    -   129. The method of any prior Aspect, wherein the spacing between        the spacers is approximately the minimum dimension of an        analyte.    -   130. The method of any prior Aspect, wherein one or both plate        sample contact surfaces comprises one or a plurality of        amplification sites that are each capable of amplifying a signal        from the analyte or a label of the analyte when the analyte or        label is within 500 nm from an amplification site.    -   131. The method of any prior Aspect, wherein the sample is        exhale breath condensate.    -   132. The method of any prior Aspect, wherein the sample is a        vapor from a biological sample, an environmental sample, a        chemical sample, or clinical sample.    -   133. The method of any prior Aspect, wherein the analyte        comprises a molecule (e.g., a protein, peptides, DNA, RNA,        nucleic acid, or other molecules), cells, tissues, viruses, and        nanoparticles with different shapes.    -   134. The method of any prior Aspect, wherein the analyte        comprises volatile organic compounds (VOCs).    -   135. The method of any prior Aspect, wherein the analyte        comprises nitrogen, oxygen, CO2, H2O, and inert gases.    -   136. The method of any prior Aspect, wherein the analyte is        stained.    -   137. The method of any prior Aspect, wherein on one of the        sample surface, it further comprises an enclosure-spacer that        encloses a partial or entire VC samples deposited on the        collection plate.    -   138. The method of any prior Aspect, wherein the highly uniform        thickness has a value equal to or less than 0.5 um.    -   139. The method of any prior Aspect, wherein the highly uniform        thickness has a value in the range of 0.5 um to 1 um.    -   140. The method of any prior Aspect, wherein the highly uniform        thickness has a value in the range of 1 um to 2 um.    -   141. The method of any prior Aspect, wherein the highly uniform        thickness has a value in the range of 2 um to 10 um.    -   142. The method of any prior Aspect, wherein the highly uniform        thickness has a value in the range of 10 um to 20 um.    -   143. The method of any prior Aspect, wherein the highly uniform        thickness has a value in the range of 20 um to 30 um.

Additional Notes

Further examples of inventive subject matter according to the presentdisclosure are described in the following enumerated paragraphs.

It must be noted that as used herein and in the appended claims, thesingular forms “a”, “an”, and “the” include plural referents unless thecontext clearly dictates otherwise, e.g., when the word “single” isused. For example, reference to “an analyte” includes a single analyteand multiple analytes, reference to “a capture agent” includes a singlecapture agent and multiple capture agents, reference to “a detectionagent” includes a single detection agent and multiple detection agents,and reference to “an agent” includes a single agent and multiple agents.As used herein, the terms “adapted” and “configured” mean that theelement, component, or other subject matter is designed and/or intendedto perform a given function. Thus, the use of the terms “adapted” and“configured” should not be construed to mean that a given element,component, or other subject matter is simply “capable of” performing agiven function. Similarly, subject matter that is recited as beingconfigured to perform a particular function may additionally oralternatively be described as being operative to perform that function.

As used herein, the phrase, “for example,” the phrase, “as an example,”and/or simply the terms “example” and “exemplary” when used withreference to one or more components, features, details, structures,embodiments, and/or methods according to the present disclosure, areintended to convey that the described component, feature, detail,structure, embodiment, and/or method is an illustrative, non-exclusiveexample of components, features, details, structures, embodiments,and/or methods according to the present disclosure. Thus, the describedcomponent, feature, detail, structure, embodiment, and/or method is notintended to be limiting, required, or exclusive/exhaustive; and othercomponents, features, details, structures, embodiments, and/or methods,including structurally and/or functionally similar and/or equivalentcomponents, features, details, structures, embodiments, and/or methods,are also within the scope of the present disclosure.

As used herein, the phrases “at least one of” and “one or more of,” inreference to a list of more than one entity, means any one or more ofthe entity in the list of entity, and is not limited to at least one ofeach and every entity specifically listed within the list of entity. Forexample, “at least one of A and B” (or, equivalently, “at least one of Aor B,” or, equivalently, “at least one of A and/or B”) may refer to Aalone, B alone, or the combination of A and B.

As used herein, the term “and/or” placed between a first entity and asecond entity means one of (1) the first entity, (2) the second entity,and (3) the first entity and the second entity. Multiple entity listedwith “and/or” should be construed in the same manner, i.e., “one ormore” of the entity so conjoined. Other entity may optionally be presentother than the entity specifically identified by the “and/or” clause,whether related or unrelated to those entities specifically identified.

Where numerical ranges are mentioned herein, the invention includesembodiments in which the endpoints are included, embodiments in whichboth endpoints are excluded, and embodiments in which one endpoint isincluded and the other is excluded. It should be assumed that bothendpoints are included unless indicated otherwise. Furthermore, unlessotherwise indicated or otherwise evident from the context andunderstanding of one of ordinary skill in the art.

In the event that any patents, patent applications, or other referencesare incorporated by reference herein and (1) define a term in a mannerthat is inconsistent with and/or (2) are otherwise inconsistent with,either the non-incorporated portion of the present disclosure or any ofthe other incorporated references, the non-incorporated portion of thepresent disclosure shall control, and the term or incorporateddisclosure therein shall only control with respect to the reference inwhich the term is defined and/or the incorporated disclosure was presentoriginally.

We claim:
 1. A device for collecting and analyzing vapor condensate (VC)sample, comprising: a. a collection plate and a cover plate, wherein:the plates are movable relative to each other into differentconfigurations;  i. one or both plates are flexible; and  ii. each ofthe plates has, on its inner respective surface, a sample contact areafor contacting a vapor condensate (VC) sample that contains an analyte; iii. wherein one of the configurations is an open configuration, inwhich: the two plates are either completely or partially separatedapart, and the VC sample is deposited on one or both of the plates; andwherein another of the configurations is a closed configuration which isconfigured after the VC sample deposition in the open configuration; andin the closed configuration: at least a part of the VC sample is betweenthe two plates and in contact with the two plates, and has a thicknessthat (a) is regulated by the two sample contact surfaces of the plateswithout using spacers, and (b) is equal to or less than 30 um with asmall variation.
 2. A system is provided herein for rapidly analyzing avapor condensate sample using a mobile phone comprising: a. a device ofclaim 1; b. a mobile communication device comprising: i. one or aplurality of cameras for the detecting and/or imaging the vaporcondensate sample; and ii. electronics, signal processors, hardware andsoftware for receiving and/or processing the detected signal and/or theimage of the vapor condensate sample and for remote communication.
 3. Amethod is provided herein for analyzing an analyte in a vapor condensatesample comprising: a. obtaining the device of claim 1; b. depositing thevapor condensate sample onto one or both pates of the device; c. placingthe plates in a closed configuration and applying an external force overat least part of the plates; and d. analyzing the analyts in the layerof uniform thickness while the plates are the closed configuration.